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Table of Contents

PIRSF Search Help

iProClass Database

iProLINK Help

Text Search Help

Text Search Result Help

Batch Retrieval Help

BLAST Search Help

BLAST Result Help

FASTA Search Help

FASTA Result Help

Related Sequence Help

Peptide Match Search

Pattern Search

Multiple Alignment Help

Pairwise Alignment Help

ID Mapping Help

Composition/Molecular Weight Calculation Help

PIRSF scan

Amino Acid Sequence Code Table

UniProtKB Identifiers for BLAST/FASTA Search and Analysis Tools Tools

iProClass Report

PIRSF Report

Master Protein Directory Help

Complete Predicted Proteomes Help

GO Slim Analysis

Master Reagent Directory Help

Representative Proteomes Help

 

PIRSF Database


What is PIRSF?
The PIRSF protein classification system is a network with multiple levels of sequence diversity from superfamilies to subfamilies that reflects the evolutionary relationship of full-length proteins and domains. The primary PIRSF classification unit is the homeomorphic family, whose members are both homologous (evolved from a common ancestor) and homeomorphic (sharing full-length sequence similarity and a common domain architecture). Automatically generated protein clusters are manually curated for membership, domain architecture, annotation of sequence features, and specific biological functions and biochemical activities, when possible.



What in PIRSF for me?
PIRSF offers curated protein families with rules for functional site and protein name propagation and standardization, therefore, improving the sensitivity of protein identification and functional inference. Searching your protein sequence against PIRSF database provides a faster and more accurate assessment of its function than a BLAST search against an uncurated protein database. It avoids pitfalls such as numerous erroneous annotations, best hits based on a domain secondary to the main protein function, spurious hits, etc.

PIRSF Definitions

-Curation Status
  • Uncurated: Computer-generated protein clusters, no manual curation. The clusters are computationally defined using both pairwise based parameters (% sequence identity, sequence length ratio and overlap length ratio) and cluster-based parameters (% matched members, distance to neighboring clusters and overall domain arrangement).

  • Preliminary: Computer-generated clusters are manually curated for membership (do proteins belong to the assigned cluster?) and domain architecture (Pfam domains listed from N- to C- termini).

  • Full/Full (with description): A name is assigned to the protein family, and accompanying references are listed when available. In many cases, brief descriptions are also provided.


  • -PIRSF Membership
  • Full (F): proteins sharing end-to-end sequence similarity and common domain architecture.

  • Associate (A): members whose lengths are outside the family length range, including sequences fragments, alternate splice and alternate initiator variants, and peptides derived from proteolytic processing, are classified as associate members with the conceptual complete sequence from which they are derived. Associate members also include individual proteins with atypical domain architecture (thus, not yet forming a separate subfamily).

  • Seed (S): full members that are used to generate family specific full-length and domain HMMs.

    Use the following formats to perform a search in iProClass using PIRSF Membership in the search field: PIRSFxxxxxx:F, PIRSFxxxxxx:S or PIRSFxxxxxx:A for Full, Seed or Associate, respectively, with xxxxxx being the PIRSF number.


  • -PIRSF Name Evidence Tag
  • [Validated]: to indicate that at least one member in the family has experimentally-validated function.

  • [Predicted]: for families whose functions are inferred computationally based on sequence similarity and/or functional associative analysis.

  • [Tentative]: cases where experimental evidence is not decisive.


  • -PIRSF Family Level
  • Homeomorphic family (HFam): members belonging to the family are both homologous (evolved from a common ancestor) and homeomorphic (sharing full-length sequence similarity and a common domain architecture). The homeomorphic family level is the primary PIRSF curation level – and most significant in terms of annotation and most invested with the biological meaning. A protein may be assigned to one and only one homeomorphic family, which may have zero or more parent nodes and zero or more child nodes.Homeomorphic families are assigned numbers < 500000, e.g., PIRSF001830

  • Subfamily (SubFam): The subfamily level is used to delineate protein clusters within a homeomorphic family that have specialized functions and/or variable domain architectures. Like its parent, each subfamily is also homologous and homeomorphic. A protein may be assigned to zero or one subfamily, which will have exactly one parent node. Subfamilies are assigned numbers ≥ 500000 and < 800000, e.g., PIRSF500000.

  • Superfamily (SuperFam): The superfamily level is used to bring together a number of distantly related families and orphan proteins that share one or more domains. Depending on the extent of domain coverage, a superfamily may be a “homeomorphic superfamily” (common domain architecture with full-length sequence coverage) or a “domain superfamily” (partial sequence coverage). Superfamilies are assigned numbers ≥ 800000, e.g., PIRSF800000.


  • -Domain Architecture
    Pfam domains assigned with high confidence, either manually or automatically (for single domain-containing families), listed from N- to C- termini. PFam domains are separated by semi-colons, although in a few cases, domains are separated by a dash indicating the presence of inserted domains. Numbers in parenthesis indicate the repetition of a domain. There is a particular syntax for this feature. For example, PF11111(1-3) allows for 1 to 3 copies of PF11111, whereas PF11111(2-) allows for any number of domains above 1 (2 or more).However, PF11111(0,2) allows for none or two copies of this domain.

    -Representative Sequence
    It is the sequence of a member of a PIRSF that belongs to a model organism (if present), belongs to UniProtKB/Swiss-Prot (if any) and meets the PIRSF criteria of uniform length and common domain architecture. Representative sequences are assigned automatically, but curator may decide to change it.



    iProClass Database



    What is iProClass?
    iProClass provides summary descriptions of protein family, function and structure for UniProt sequences, with links to over 90 biological databases (See data sources). iProClass comprises reports for all UniProtKB proteins and those proteins that are exclusively in UniParc database.

    iProClass Text Search
    Retrieve a matching list of summary reports by text string or unique identifier (selecting a field from the dropdown menu). Click "Search" button to retrieve results. You may open extra input boxes in your query by clicking on "Add input box". See Text Search Help for additional information.





    iProLINK Help



    What is iProLINK?
    As PIR focuses its effort on the curation of the UniProtKB protein sequence database, the goal of iProLINK is to provide curated data sources that can be utilized for text mining research in the areas of bibliography mapping, annotation extraction, protein named entity recognition, and protein ontology development. The process of applying literature mining methods for protein database curation involves several tasks:
    • Bibliography Mapping: identification of articles from literature sources (such as PubMed) that describe a given protein entry;
    • Annotation Extraction: categorization of annotation types and extraction of sentences and/or phrases describing the given annotation; and
    • Database Curation: conversion of the extracted literature information into annotation in the database with structured syntax, controlled vocabulary, and evidence attribution.

    These tasks are also related to the topics of protein named entity recognition and protein ontology development. A prerequisite to bibliography mapping is protein named entity recognition/identification of protein names from articles. Furthermore, due to the long-standing problem of protein nomenclature, a protein ontology can assist entity recognition with the description of names and synonyms of protein classes as well as their relationships.

    Bibliography Mapping
    Linking protein entries to relevant scientific literature that describes or characterizes the proteins is crucial for increasing the amount of experimentally verified data and for improving the quality of protein annotation.
    Enter a text string or a unique identifier to retrieve bibliography related to you query.
    Results are shown in a table similar to iProClass result page, but with specific columns related to bibliography retrieval. Links to the bibliography record as well as the individual PubMed entries are available.
    iProLINK Bibliography Mapping Result Page for UniProtKB P13866



    Feature Evidence Attribution
    In the PIR-PSD database, feature annotation such as binding sites, catalytic sites, and modified sites are labeled with status tags "experimental" or "predicted" to distinguish experimentally verified from computationally predicted data. To appropriately attribute bibliographic data to features with experimental evidence, a retrospective literature survey was conducted, which involves both citation mapping (finding citations from the Reference section that describe the given experimental feature) and evidence tagging (tagging the sentences providing experimental evidence in an abstract and/or full-text article). Now this is being extended to UniProtKB proteins. Search for proteins with a particular feature attribution such as post- translational modifications, or enter a UniProtKB identifier to retrieve feature attribution for your query. Later on you may get curated bibliography related to the entry by clicking on "Bibliography" or all available tagged evidence by clicking on "Tagged Evidence"


    Examples for Feature Evidence Attribution Text Search

    Feature Example
    Active siteser
    Binding siteheme
    Cleavage sitearg
    Cross-linkcys
    Disulfide bondsall
    Modified siteblock*
    Domainsignal sequence
    Productdermorphin
    RegionATP binding



    iProLINK Feature Mapping Result Page for heme binding site



    RLIMS-P

    The RLIMS-P is a rule-based text-mining program specifically designed to extract protein phosphorylation information on protein kinase, substrate and phosphorylation sites from the abstracts (Hu et al., 2005). Submission of PMIDs as input, returns a summary table for all PMIDs with links to full reports . The summary table lists the PMID of each phosphorylation-related abstract along with its top-ranking annotation result, followed by a list of remaining PMIDs for abstracts containing no phosphorylation information. Full reports can be retrieved from the summary table using hypertext links (from “text evidence") or by selecting one or more PMID(s) in the list.

    Summary Table for PMIDs 2108025, 16436437, 15193450



    The full RLIMS-P report contains five sections (see table below): 1- PubMed citation information (publication date, authors, journal); 2- PMID mapping to UniProtKB, consisting of the accession, ID, protein name, organism, and protein family of the mapped entry, with links to UniProtKB and iProClass (Wu et al., 2004) protein reports containing rich biological and functional information; 3- Name mapping to UniProtKB, including options to use either names appeared in the abstract or user-specified names for searching online BioThesaurus; 4- Annotation with a ranked list of RLIMS-P extraction results for each set of the three phosphorylation objects; and 5- Text evidence showing the original abstract and title, with extracted objects tagged in different colors to distinguish protein kinases, phosphorylated proteins, and phosphorylated residues/positions. An option is provided for turning on/off the color-tagging for each type of object in the abstract for visual inspection.

    Full Report for PMID 2108025




    Entity Recognition/Ontology Development
    Protein named entity recognition (finding protein names from literature texts) is a prerequisite for bibliography mapping (identifying papers describing specified proteins). It is also fundamental for several other biological literature mining tasks, including the extraction of protein annotations (such as protein-protein interactions) from literature.

    -BioThesaurus
    BioThesaurus is a web-based system designed to map a comprehensive collection of protein and gene names to UniProtKB protein entries. Currently covering more than two million proteins, BioThesaurus consists of over 2.6 million names extracted from multiple online resources based on database cross-references in iProClass. It allows the retrieval of synonymous names of given protein entries and the identification of ambiguous names shared by multiple proteins.
    You can search by typing a gene/protein name, or alternatively, by using an identifier.

    BioThesaurus Report for UniProtKB P18688





    -Protein Name Dictionary and Word Token Dictionaries
    PIR protein name dictionary is derived from the protein name field in the iProClass database, which consists of protein names from UniProt (Swiss-Prot,TrEMBL, PIR-PSD) and RefSeq. After the initial compilation, the dictionary undergoes several filtering processes to generate unique protein names (including synonyms and acronyms), and to remove nonsensical names and certain general descriptional annotations. For example, entry names such as "Inter-alpha-trypsin inhibitor (GIK-14) (Fragment)" were broken into Inter-alpha-trypsin inhibitor, GIK-14 and Fragment. The name Fragment is later removed from the dictionary along with a list of other "bad" names such as hypothetical protein, conserved hypothetical protein, unnamed protein product, predicted protein, and predicted protein of unknown function. In addition, words such as probable, putative, and similar to before protein names are also removed so that a name like putative aspartate aminotransferase A is merged to aspartate aminotransferase A to reduce the redundancy. Derived from over 1.5 million iProClass entries, the protein name dictionary currently has about 700,000 names, each of which is shown with its frequency count. Most protein names are composed of combinations of two or more words (or tokens). Therefore, protein name rules can be derived from tokenized protein words and used during post-tagging processing to improve machine learning-based named entity recognition. We have compiled specialized single-word dictionaries by tokenization and classification of protein names from 30,000 well-curated iProClass protein entries (each containing at least 5 reference citations). The dictionaries consist of individual word tokens categorized into five classes.

    • Biomedical Terms (bt): These terms are used in a broad of biological and medical sciences. They mainly describe structures of all forms of life at different levels (from gross morphology to molecular structure), as well as their respective functions and mechanisms in both normal (physiological) and diseased states (pathological).
    • Chemical Terms (ct): These are words that describe organic or inorganic chemical materials, chemical groups or bonds, or chemical properties.
    • Macromolecules (mc): These words refer to biopolymers such as proteins, peptides, DNA, RNA, polysaccharides, or glycoproteins.
    • Common English (ce): Common English words are used to describe various aspects or properties of proteins, such as short, signal, interacting, and repair. These also include spelled-out forms of Greek letters, as well as stop words like of, at, and to.
    • Non-word tokens: They are combinations of letters, numbers, or symbols. They often are acronyms, synonyms, or abbreviations. The form of non-word tokens can be number only, single letter, multiple letters, or combinations of numbers, letters, and other symbols. Non-word tokens may stand for biochemical entities such as nucleic acids, nucleotide, and amino acids, which can be used in a protein name.


    -Protein Name Tagging Guidelines and Name-Tagged Corpora
    Other iProLINK data resources for named entity recognition are two sets of literature corpora that were manually tagged with protein names based on two versions of tagging guidelines. Guideline 1.0 defines how to tag protein objects, not protein named entities, whereas Guideline 2.0 defines tagging rules for protein named entities regardless of the context of the object.


    -PIRSF Family Classification-Based Protein Ontology
    Biological ontologies are crucial for biological knowledge management, including mining literature data to extract relevant information and integrating information from multiple databases. A protein ontology?consisting of names and synonyms of protein classes as well as their relationships?can be used to assist with protein named entity recognition. Furthermore, an ontology based on protein family relationships, such as the PIRSF classification system, can be mapped to and complement the Gene Ontology (GO). We have developed a protein ontology based on PIRSF hierarchical family names. The ontology is in the GO flat file format with a DAG (directed acyclic graph) structure, and can be browsed using application tools such as DAG-Edit.





    Text Search Help



    Select a Database
    Depending on the route by which the Text Search page was accessed, you may need to select between the iProClass database, which includes UniProtKB and unique UniParc proteins, and PIRSF database, which includes the whole set of PIRSF families (i.e., any curation level). However, Text Search in the home page and the quick search box uses iProClass database.
    The output file will display a table with individual protein entries or protein families, for iProClass or PIRSF, respectively.

    Selecting a Field
    Searchable fields are selected from the dropdown menu. The items will vary according to database selected, since each database contains different types of information.

    Query Input
    Enter a unique identifier or other search string in the box provided.
    Certain items (such as "Length" or anything with "ID") will be exact match searches. Other items will be substring searches (as if preceded and followed by wild cards).
    When the field option is present, entering "not null" in the text box will cause the search to return only those entries that have some data in the selected field, while entering "null" will return only those that lack data in the selected field.
    For peptide search, type in a string of amino acid residues in single letter code (at least three letters), then press the arrow to retrieve results.

    Add Input Box Button
    If desired, multiple fields can be searched simultaneously. Pressing the "+ box" button adds another query line, up to a maximum of 8 lines. The added input boxes are connected by logical operator choices (see below), with the default being the Add operator.

    Using the Logical Operators AND, OR, NOT
    Search supports the logical 'AND', 'OR', and 'NOT' operators. For example, to retrieve results that include Pfam domains A or B, type A in your first query field and add a query line by clicking the "Add input box" button. Enter your 2nd query (B) and select the OR operator. Similarly, to retrieve multi-domain proteins that have both Pfam domains A and B, use the 'AND' operator. Proteins that have domain A and not domain B can be retrieved using the 'NOT' operator.

    Number of Results
    To provide the fastest result, the default number of entries shown on any one page is 50.

    Search Categorizations and Unique Identifiers
    The following table indicates the searchable fields for Text and Batch Retrieval search functions. Searchable fields can be selected from the dropdown menu. You can search the databases using unique identifiers or keywords. Entry examples are shown below.

    MAIN CATEGORY SUBCATEGORY INCLUDED FIELDS EXAMPLE(S) iProClass PIRSF
    SequenceUniProt IDsUniProtKB ID/ACIMDH2_HUMAN/P12268
    UniRef100UniRef100_P12268
    UniParcUPI00004C7276
    PIR Sequence IDsPIR-PSD ID/ACA31997/I52303
    Other Sequence IDs
    FlyBase IDFBgn0002940
    GenBank ACJ04208
    GenPept ACAAH12840.1
    NCBI GI Number15277480
    IPI IDIPI00291510
    MGI ID95561
    RefSeq ACNM_000875
    SGD IDS000001047
    TIGR IDSAG1156
    Gene/Protein NameGene NameIMPDH2
    Protein NameIMP dehydrogenase
    ClassificationClass IDs
    BLOCKS IDIPB000644
    COG IDCOG1009
    Pfam ID/NamePF00478
    IMP dehydrogenase
    PIRSF ID/NamePIRSF000130
    inosine-5'-monophosphate dehydrogenase
    PRINTS IDPR01434
    PROSITE IDPS00487
    SCOP Superfamily NameInosine monophosphate dehydrogenase
    UniRef50 IDUniRef50_P20839
    UniRef90 IDUniRef90_P12268
    PIRSFAverage Length500 (amino acids)
    Curation StatusFull
    PIRSF MembershipPIRSF000186:F
    PIRSF LevelHFam
    Rep. Seq.P12268
    FunctionComplex/InteractionsBIND ID93185
    Gene OntologyGO ID0003938
    Pathway KEGG Pathway/IDPurine Metabolism
    hsa00230
    EnzymeEC Number/NameEC 1.1.1.205/ Oxidoreductases
    FeatureRESID IDAA0005
    All Feature FieldsL-cysteine
    OrganismTaxonomyTaxon
    Group/Group ID
    Euk/Mammal / 40674
    Taxon ID9606
    Species/LineageLineageEukaryota
    Organism NameHomo sapiens
    Common NameHuman
    LiteraturePubMed IDPubMed ID10097070
    Author NameAuthor NameHuberman
    Journal NameJournal NameBiochem Biophys Res Commun.
    Paper TitlePaper TitleCloning and sequence of the human type II IMP
    Miscellaneous 3D StructurePDB ID1B3O
    PropertyLength514 (amino acids)
    Molecular Weight55804 (Daltons)
    KeywordKeywordNAD
    Genetic Variation/DiseaseOMIM ID146691
    Genome Entrez Gene ID3615







    Text Search Result Help

    This section describes the Text Search Result page for iProClass and PIRSF. The overall layout of the page is similar for both, however, it differs in the default columns that are displayed, as well as the tools available to analyze data. Common features are described first 1-3, where numbers indicate the place where you will find these features in the page (see iProClass and PIRSF result pages below).

    1- Search
    This feature allows you to perform an additional search in case you want to further filter out your output or you want to start a new search (no need to go back to the previous page unless you want to use a different database).

    2- Display Options
    Depending on your specific need you can choose the columns to be displayed. To do this, click on the "Display Option" button, select the relevant field(s) in the "Fields Not in Display" list and transfer them to the "Fields in Display" list via the ">" button. Conversely, columns can be removed from display. Finally, click on "apply" for the changes to take effect.

    3- Save Results As
    The output can be saved to the user's local computer. The results will be saved for selected entries or, if no proteins are selected, for all entries. Clicking "Table" will save the displayed columns as a tab-delimited text file, which may be imported into a spreadsheet for easier viewing or analysis. Clicking "FASTA" will save the IDs and sequences in FASTA format.

    iProClass Result Page



    4- Analyze: BLAST, FASTA, Pattern Match, Multiple Alignment and Domain Display
    Retrieved entries can be further analyzed using the sequence analysis programs available in the Results page. First, select the protein(s) using the checkboxes on the left side of the table, then click the corresponding analysis tool.
  • Click "BLAST" or "FASTA" button, and a new query page will be displayed, along with the parameters that were selected in the initial search.
  • Click "Pattern Match" to search against the PROSITE database.
  • For multiple alignment, check at least 2 proteins (but no more than 70), then click the "Multiple Alignment" button. This will open the Multiple alignment form from which you can select one of the following alignment programs: ClustalW, T-Coffee or Muscle. The result page will display the alignment and alignment viewer. For ClustalW and T-Coffee, the neighbor-joining tree and alignment can be viewed, edited and saved using either PIR-TAV viewer or JalView. For Muscle only Jalview is available.
  • Domain display option, shows PFam domains (if present) in graphical format.


  • 5- Results Display
    Results of the search are displayed in a customizable table. The exact columns displayed will depend on the fields searched for, and user preference.

    Protein AC/ID
    The Protein AC/ID refers to the UniProtKB or UniParc identifiers. Below these numbers, you may choose either the iProClass or the UniProtKB/UniParc view of the protein report. The source of the UniProtKB sequence is shown as UniProtKB/Swiss-Prot or UniProtKB/TrEMBL if the protein sequence is from Swiss-Prot or TrEMBL section, respectively. Alternatively, the UniParc ID will be displayed if the sequence is no present in the UniProtKB database along with the UniParc report.

    Protein Name
    The common name given to a protein, that identifies its function or specifies its features.

    Length
    Number of amino acid residues in the peptide or protein.

    Organism Name
    The genus and species of the source organism from which the sequence originated. Links to NCBI taxonomy information are provided.

    PIRSF ID
    If a protein belongs to a PIRSF family, then this column will display the corresponding family identifier. Click on the ID to retrieve the PIRSF report (see annotated output).

    Related Seq.
    This column shows the number of pre-computed BLAST hits obtained using default parameters. Only up to 300 sequences will be displayed. By clicking the number you access to the related sequence page. This allows to have a glance at sequence similarity in a very fast way. The number itself already provides some information about how unique the protein is. For example, a very low number may tell you that the query is specific to a certain species, genus, taxon, etc. The "+" sign next to the Related Sequence title allows to compare number of related sequences at 3 different E-value cut-offs as shown below.



    Matched Fields
    The field(s) matched by the query.

    6- GO Slim
    GO slims are smaller versions of the Gene Ontologies containing a subset of the terms in the whole GO. They give a broad overview of the ontology content without the detail of the specific fine grained terms. GO slims are particularly useful for giving a summary of the results of GO annotation of a genome or proteome when broad classification of gene product function is required.

    You can view the GO slim terms for Biological Function, Component, and Process by selecting the “Show GO Slim” button in the analysis tool bar A. You can then view statistics for the individual ontologies (Function, Component, and Function) by checking entries of interest and selecting the ontology to show (for example, function in this example, A). Follow the GO ID links to learn more about the GO term. In addition, you can view the terms within the ontology by selecting the GO graphical hierarchical view icon (B). The graphical display will show the GO hierarchy in relation to the terms shown in the table C). Terms that map to a given protein are shown in color, the number in parenthesis indicates the number of proteins in the selected set that are annotated with the given term. Clicking on the number will retrieve the corresponding protein entries. An option to display the graph in svg format is provided. This format allows to rescale the image.



    PIRSF Result Page



    4- Analyze: Multiple Alignment, Taxonomic Distribution and Domain Display
    Retrieved entries can be further analyzed using the sequence analysis programs available in the Results page. First, select the PIRSF(s) using the checkboxes on the left side of the table, then click the corresponding analysis tool.
  • Click on the "Multiple Alignment" button to generate an alignment and neighbor-joining tree for selected PIRSFs. If only one PIRSF is selected, then alignment can be performed using the seed members or the full set of sequences (see below). Alternatively, you can modify the list, adding/deleting members. Please note that there is a maximum of 70 sequences for the alignment.



    If more than one PIRSF is selected, a pop-up message will indicate that the representative sequences of each family will be used for the alignment.



    See sample output for selected PIRSF000186 members.
  • Click on Taxonomic Distribution to look at the number of members of a PIRSF present in the different taxa. You can perform this function on one or multiple PIRSFs. If only one PIRSF is selected, the taxonomic distribution for the parent family and the children (if any) will be displayed. See sample output for PIRSF000186.
  • Domain display option, shows PFam domains (if present) in a graphical format.


  • 5- Results Display
    The results of the search are displayed in a customizable table. The exact columns displayed will depend on the fields searched for, and user preference.

    PIRSF ID
    This column will display the corresponding family identifier. Click on the ID to retrieve the PIRSF report. The icon below the PIRSF ID states the family level, namely homeomorphic family (HFam), subfamily (SubFam), and superfamily (SuperFam). Clicking on this icon will bring the DAG view of PIRSF Hierarchy with the PFam domain at the higher level.

    PIRSF Name
    PIRSF name is meaningful for fully curated families since these names have been analyzed by a curator. The PIRSF Name aims at describing some common characteristic or function of the members in the family. Whenever possible PIRSF names are based on literature and try to follow any published standards. In most cases, names have an evidence tag or name status attached in brackets, which provides the level of confidence on the assigned name. The possible PIRSF name status tags are: validated (there is experimental data supporting PIRSF name), tentative (there is no conclusive experimental evidence) or predicted (predicted by computational methods).

    Num. of Seq.
    Number of sequences in the family.

    Av.Length
    It is the protein average length of the members of the PIRSF.

    Domain Architecture
    It represents the curated information regarding present Pfam domains in the protein family. Pfam domains are listed in order from N- to C- terminus separated by semi-colons. In a few cases, domains are separated by a dash indicating the presence of inserted domains. Numbers in parenthesis indicate the repetition of a domain. There is a particular syntax for this feature. For example, PF11111(1-3) allows for 1 to 3 copies of PF11111, whereas PF11111(2-) allows for any number of domains above 1 (2 or more). However, PF11111(0,2) allows for none or two copies of this domain.

    Curation Status
    Uncurated: Computer-generated protein clusters, not curated.
    Preliminary: Membership and domain architecture of protein families determined by manual curation.
    Full:Protein family name with accompanying references (when available), and sometimes brief descriptions, provided after thorough manual curation.

    Matched Fields
    The field(s) matched by the query.




    Batch Retrieval Help


    Retrieve multiple entries by selecting a specific identifier or a combination of identifiers.

    Select the database
    Before entering the protein identifiers you need to select the most convenient database. If you want to retrieve information about protein families, then select the PIRSF database, however if you are interested in analyzing individual proteins, then select the iProClass database.

    Rules for entering IDs
    Multiple Entry IDs should be separated by lines or spaces.
    IDs may be specified as a single category or as mixed categories. However, if your entries have the same type of ID, it is recommended that you define the ID field to speed up the retrieval process.


    You can further analyze and save any retrieved results and/or their respective sequences in a similar way as you do after a Text Search. Click on "Show Match List" to check the correspondence between your entry identifiers and the ones from the selected database.

    Batch Retrieval Output in iProClass Database

    Retrieval of sequences with GI numbers 1169968, 1707983 and 304131



    1- Retrieval Box
    This box shows your query ID and also allows you to perform a new retrieval.

    2- Display Options
    Depending on your specific need you can choose the columns to be displayed. To do this, click on the "Display Option" button, select the relevant field(s) in the "Fields Not in Display" list and transfer them to the "Fields in Display" list via the ">" button. Conversely, columns can be removed from display. Finally, click on "apply" for the changes to take effect.

    3- Save Results As
    The output can be saved to the user's local computer. The results will be saved for selected entries or, if no proteins are selected, for all entries. Clicking "Table" will save the displayed columns as a tab-delimited text file, which may be imported into a spreadsheet for easier viewing or analysis. Clicking "FASTA" will save the IDs and sequences in FASTA format.

    4- Analyze: BLAST, FASTA, Pattern Match, Multiple Alignment and Domain Display
    Retrieved entries can be further analyzed using the sequence analysis programs available in the Results page. First, select the protein(s) using the checkboxes on the left side of the table, then click the corresponding analysis tool.
  • Click "BLAST" or "FASTA" button, and a new query page will be displayed, along with the parameters that were selected in the initial search.
  • Click "Pattern Match" to search against the PROSITE database or against a user defined pattern.
  • For multiple alignment, check at least 2 proteins (but no more than 70), then click the "Multiple Alignment" button. This will open the Multiple alignment form from which you can select one of the alignment programs: ClustalW, T-Coffee or Muscle. The result page will display the alignment and alignment viewer. For ClustalW and T-Coffee, the neighbor-joining tree and alignment can be viewed, edited and saved using either PIR-TAV viewer or JalView. For Muscle only Jalview is available.
  • Domain display option, shows PFam domains (if present) in graphical format.


  • 5- Results Display
    Results of the search are displayed in a table.

    Protein AC/ID
    The Protein AC/ID refers to the UniProtKB accession number and id, respectively. Below these, you may choose either the iProClass or the UniProtKB/UniParc view of each protein report. The source of the UniProtKB sequence is shown as UniProtKB/SP or UniProtKB/Tr if the protein sequence is from Swiss-Prot or TrEMBL, respectively.

    Protein Name
    The common name given to a protein, that identifies its function or specifies its features.

    Length
    Number of amino acid residues in the peptide or protein.

    Organism Name
    The genus and species of the source organism from which the sequence originated. Links to NCBI taxonomy information are provided.

    PIRSF ID
    If a protein belongs to a PIRSF family, then this column will display the corresponding family identifier. Click on the ID to retrieve the PIRSF report.

    Match Range
    This column displays in red the peptide query within the sequence.

    6- GO Slim
    GO slims are smaller versions of the Gene Ontologies containing a subset of the terms in the whole GO. They give a broad overview of the ontology content without the detail of the specific fine grained terms. GO slims are particularly useful for giving a summary of the results of GO annotation of a genome or proteome when broad classification of gene product function is required.

    You can view the GO slim terms for Biological Function, Component, and Process by selecting the “Show GO Slim” button in the analysis tool bar. You can then view statistics for the individual ontologies (Function, Component, and Function) by checking entries of interest and selecting the ontology to show (for example, function in this example). Follow the GO ID links to learn more about the GO term.

    7- Show match list
    Shows a table mapping your query IDs with the UniProtKB/UniParc IDs.

    Batch Retrieval Output in PIRSF Database

    Retrieval of PIRSFs using as queries UniProtKB ACs P51375, P09831 and Q05755



    1- Retrieval Box
    This box shows your query ID and also allows you to perform a new retrieval

    2- Display Options
    Depending on your specific need you can choose the columns to be displayed. To do this, click on the "Display Option" button, select the relevant field(s) in the "Fields Not in Display" list and transfer them to the "Fields in Display" list via the ">" button. Conversely, columns can be removed from display. Finally, click on "apply" for the changes to take effect.

    3- Save Results As
    The output can be saved to the user's local computer. The results will be saved for selected entries or, if no proteins are selected, for all entries. Clicking "Table" will save the displayed columns as a tab-delimited text file, which may be imported into a spreadsheet for easier viewing or analysis. Clicking "FASTA" will save the IDs and sequences in FASTA format.

    4- Analyze: Multiple Alignment, Taxonomic Distribution and Domain Display
    Retrieved entries can be further analyzed using the sequence analysis programs available in the Results page. First, select the PIRSF(s) using the checkboxes on the left side of the table, then click the corresponding analysis tool.
  • Click on the "Multiple Alignment" button to generate a ClustalW alignment and neighbor-joining tree for the members of the selected PIRSFs. If one PIRSF is selected, then alignment of all its seed members will be performed (see sample output for PIRSF000186). But if many PIRSFs are selected, then alignment of the representative sequences will be displayed.
  • Click on Taxonomic Distribution to look at the number of members of a PIRSF present in the different taxa. You can perform this function on one or multiple PIRSFs. If only one PIRSF is selected, the taxonomic distribution for the parent family and the children (if any) will be displayed. See sample output for PIRSF000186.
  • Domain display option, shows PFam domains (if present) in a graphical format. See sample output or annotated output for PIRSF000186.


  • 5- Results Display
    Results of the search are displayed in a table.

    PIRSF ID
    This column will display the corresponding family identifier. Click on the ID to retrieve the PIRSF report. The icon below the PIRSF ID states the family level, namely homeomorphic family (HFam), subfamily (SubFam), and superfamily (SuperFam). Clicking on this icon will bring the DAG view of PIRSF Hierarchy with the PFam domain at the higher level.

    PIRSF Name
    The name given to the family that identifies its function or specifies its features. Depending on the curation status, this name is assigned automatically (no manual curation) or manually (full curation). If the family is curated, it may display a tag next to the name indicating its status, namely validated (there is experimental data supporting PIRSF name), tentative (there is no conclusive evidence) or predicted (predicted by computational methods).

    Length
    It is the protein average length of the PIRSF members.

    Domain Architecture
    It represents the curated information regarding present Pfam domains in the protein family. Pfam domains are listed in order from N- to C- terminus separated by semi-colons. In a few cases, domains are separated by a dash indicating the presence of inserted domains. Numbers in parenthesis indicate the repetition of a domain. There is a particular syntax for this feature. For example, PF11111(1-3) allows for 1 to 3 copies of PF11111, whereas PF11111(2-) allows for any number of domains above 1 (2 or more). However, PF11111(0,2) allows for none or two copies of this domain.

    Curation Status
    Uncurated: Computer-generated protein clusters, not curated.
    Preliminary: Membership and domain architecture of protein families determined by manual curation.
    Full:Protein family name with accompanying references (when available), and sometimes brief descriptions, provided after thorough manual curation.

    Matched Fields
    The field(s) matched by the query.

    6- Show match list
    Shows a table mapping your query IDs to the corresponding PIRSF.






    BLAST Search Help



    What is BLAST?
    The Basic Local Alignment Search Tool (BLAST) allows rapid sequence comparison that optimizes the high-scoring segment pair (HSP), a measure of local similarity. For more information visit
    NCBI-BLAST.



    1. Select a Database
    BLAST search can be performed against:
  • UniProtKB is the central hub for the collection of functional information on proteins, with accurate, consistent, and rich annotation. It consists of two sections: a section containing manually-annotated records with information extracted from literature and curator-evaluated computational analysis (UniProtKB/Swiss-Prot), and a section with computationally analyzed records that await full manual annotation (UniProtKB/TrEMBL).
  • A subset of UniProtKB entries belonging to a certain organism or taxon group.
  • UniRef100 provides clustered sets of sequences at 100 % from UniProt Knowledgebase (including splice variants and isoforms) and selected UniParc records (mainly from PDB, Refseq and Ensembl databases), in order to obtain complete coverage of sequence space while hiding redundant sequences (but not their descriptions) from view. It combines identical sequences and sub-fragments with 11 or more residues (from any organism) into a single UniRef entry, displaying the sequence of a representative protein, the accession numbers of all the merged UniProtKB entries, and links to the corresponding UniProtKB and UniParc records.


  • .
    2. BLAST Search Format
    The BLAST 'Search' input area accepts:

  • ">" followed by a UniProtKB sequence identifier. Spaces between letters in the input are not allowed, although spaces before or after the identifier are allowed.


  • Raw amino acid sequence format:
    MSEPQRLFFAIDLPAEIREQIIHWRATHFPPEAGRPVAADNLHLTLAFLGEVS
    AEKEKALSLLAGRIRQPGFTLTLDDAGQWLRSRVVWLGMRQPPRGLIQLAN
    MLRSQAARSGCFQSNRPFHPHITLLRDASEAVTIPPPGFNWSYAVTEFTLYA
    SSFARGRTRYTPLKRWALTQ

  • The query sequence intersperse with numbers and/or spaces, such as:
    1    msepqrlffa idlpaeireq iihwrathfp peagrpvaad nlhltlaflg evsaekekal
    61   sllagrirqp gftltlddag qwlrsrvvwl gmrqpprgli qlanmlrsqa arsgcfqsnr
    121  pfhphitllr daseavtipp pgfnwsyavt eftlyassfa rgrtrytplk rwaltq

  • Sequences are expected to be represented in the standard IUB/IUPAC amino acid codes, with these exceptions: lower-case letters are accepted and are mapped into upper-case; U and * are acceptable letters (see table). Numerical digits in the query sequence are automatically removed.

    BLAST Options
    BLAST is performed using the BLOSUM62 matrix with default values for gap opening and extension cost. However, the following parameters can be set:

    -Composition-based Statistics
    BLAST permits calculated E-values to take into account the amino acid composition of the individual database sequences involved in reported alignments. This improves E-value accuracy, thereby reducing the number of false positive results. The improved statistics are achieved with a scaling procedure that in effect employs a slightly different scoring system for each database sequence. As a result, raw BLAST alignment scores in general will not correspond precisely to those implied by any standard substitution matrix. Furthermore, identical alignments can receive different scores, based upon the compositions of the sequences they involve.

    -Filter
      Low-complexity
      Masks off segments of the query sequence that have low compositional complexity, as determined by the SEG program of Wootton & Federhen (Computers and Chemistry, 1993). Filtering can eliminate statistically significant but biologically uninteresting reports from the BLAST output (e.g., hits against common acidic-, basic- or proline-rich regions), leaving the more biologically interesting regions of the query sequence available for specific matching against database sequences. Filtering is only applied to the query sequence, not to database sequences.

      Mask for Lookup Table Only
      This option masks only for purposes of constructing the lookup table used by BLAST. BLAST searches consist of two phases, finding hits based upon a lookup table and then extending them. The option to "Mask for lookup table only" masks only for the lookup table so that no hits are found based upon low-complexity sequence. The BLAST extensions are performed without masking and so they can be extended through low-complexity sequence. This option is still experimental and may change in the near future.

      Mask Lower Case
      With this option selected you can cut and paste a FASTA sequence in upper case characters and denote areas you would like filtered with lower case. This allows you to customize what is filtered from the sequence during the comparison to the BLAST databases.


    -Expect
    The Expect threshold establishes a statistical significance threshold for reporting database sequence matches. The default value is 10, meaning that 10 matches are expected to be found merely by chance. Lower Expect thresholds are more stringent, leading to fewer chance matches being reported. Increasing the expected threshold shows less stringent matches and is recommended when performing searches with short sequences as a short query is more likely to occur by chance in the database than a longer one, so even a perfect match (no gaps) can have low statistical significance and may not be reported. Increasing the Expect threshold allows you to look farther down in the hit list and see matches that would normally be discarded because of low statistical significance.

    -Word Size
    The word size indicates the length of the initial sequence that must be matched between the database and the query sequence.

    -Matrix
    A key element in evaluating the quality of a pairwise sequence alignment is the "substitution matrix", which assigns a score for aligning any possible pair of residues. The matrix used in a BLAST search can be changed depending on the type of sequences you are searching with. The user may choose from a list of matrices that cover various evolutionary constraints (more information can be found in a description of BLAST scoring matrices). For each matrix, a default matrix-dependent gap cost is displayed. Gap costs are described below.

    -Matrix-dependent Gap Cost
    The pull down menu shows the Gap Costs (penalty to open gap and penalty to extend gap). There are a limited number of options for these parameters. Increasing the Gap Costs will result in alignments that decrease the number of Gaps introduced. The gap open penalty is the score taken away for the initiation of a gap in a sequence. To make the match more significant the user can try making the gap penalty larger. The gap extension penalty is added to the gap open penalty for each residue in the gap, effectively penalizing longer gaps. If the user does not like long gaps, they can increase the extension gap penalty. Usually one would expect a few long gaps rather than many short gaps, so the gap extension penalty should be lower than the gap penalty. An exception is where one or both sequences are single reads with possible sequencing errors, in which case you would expect many single base gaps. The user can get this result by setting the gap open penalty to zero (or very low) and using the gap extension penalty to control gap scoring.

    -Adjust Gap Costs
    Alignments between sequences are often optimized by allowing gaps within one or both sequences. Like mismatches between aligned residues, gaps have a "cost" associated with them. There are separate penalties to open and to extend gaps. Increasing the Gap Costs will result in alignments that decrease the number and size of Gaps introduced. The Gap Open cost (or Gap Existence cost) is the score taken away for the initiation of a gap in a sequence. To make the match more significant the user can try making this gap penalty larger. The Gap Extend cost is added to the Gap Open cost for each residue in the gap, effectively penalizing longer gaps. The user can therefore select against long gaps by increasing this penalty. Usually one would expect a few long gaps rather than many short gaps, so the Gap Extend cost should be lower than the Gap Open cost. The Gap Costs can be adjusted relative to the default value using the pull down menu.

    -Number of Hits to Display
    Restricts the number of BLAST hits of matching sequences that will be reported.

    -Alignment
    Aligns your query sequence and database matches in pairs. Matches are connected with a "|" symbol. Mismatches are opposed with a space. Gaps are introduced with a "-" symbol.

    References
      Wootton JC, and Federhen S (1993) Statistics of local complexity in amino acid sequences and sequence databases. Computers and Chemistry 17:149-163.





    BLAST Results Help

    A sample output for a BLAST search against the UniProtKB database is shown below



    1- Query Sequence on
    Click on this button to display the query sequence. Conversely, click it again to hide it.

    2- Save Results As
    Search results can be saved to the user's local computer. The results will be saved for selected entries or, if no proteins are selected, for all entries. Clicking "Table" will save the displayed columns as a tab-delimited text file, which may be imported into a spreadsheet for easier viewing or analysis. Clicking "FASTA" will save the IDs and sequences in FASTA format.

    3- BLAST, FASTA, Pattern Match, Multiple Alignment and Domain Display
    Retrieved entries can be further analyzed using the sequence analysis programs available in the Results page. First, select the protein(s) using the checkboxes on the left side of the table, then click the corresponding analysis tool.
  • Click "BLAST" or "FASTA" button, and a new query page will be displayed, along with the parameters that were selected in the initial search.
  • Click "Pattern Match" to search against the PROSITE database.
  • For multiple alignment, check at least 2 proteins (but no more than 70), then click the "Multiple Alignment" button. This will open the Multiple alignment form from which you can select one of the alignment programs: ClustalW, T-Coffee or Muscle. The result page will display the alignment and alignment viewer. For ClustalW and T-Coffee, the neighbor-joining tree and alignment can be viewed, edited and saved using either PIR-TAV viewer or JalView. For Muscle only Jalview is available.
  • Domain display option, shows PFam domains (if present) in graphical format.


  • 4- Results Display

    Sort Columns
    Columns can be sorted by the corresponding values by clicking on the arrow next to the column title. By default the table is sorted by Score.

    The results from the search are displayed in a table with the following default columns:

    Protein AC/ID
    The Protein AC/ID refers to the UniProtKB accession number and ID, respectively. Below these identifiers, you may choose either the iProClass or the UniProtKB view of the protein report. The source of the UniProtKB sequence is shown as UniProtKB/Swiss-Prot or UniProtKB/TrEMBL if the protein sequence is from the Swiss-Prot or TrEMBL section of UniProtKB, respectively.

    Protein Name
    The common name given to a protein, that identifies its function or specifies its features.

    Length
    Number of amino acid residues in the peptide or protein.

    Organism Name
    The genus and species of the source organism from which the sequence originated. Links to NCBI taxonomy information are provided.

    PIRSF ID
    If a protein belongs to a PIRSF family, then this column will display the corresponding family identifier. Click on the ID to retrieve the PIRSF report (see annotated output).

    BLAST Sequence Similarity Columns
    There are three columns related to BLAST results. The Alignment column, in the extreme right, shows the BLAST results in graphical format. The top bar represents the query sequence. The bars below show the region(s) on other sequences matched to the query sequence. The bar color indicates the magnitude of the BLAST score. Click on one of these bars to see its BLASTalignment paired with the query sequence.

    SSearch Columns
    SSearch is a pairwise implementation of the Smith-Waterman alignment algorithm. When two sequences are aligned, only the shared region is shown. Within the shared region, amino acid residues from one or both sequences can be aligned with either amino acids or gaps from the other sequence. The total length of the shared region, including gaps, is represented under the Overlap column. The percentage of identical residues in the alignment is given under %iden. Clicking on that number displays the SSearch full-length alignment.

    5- Show Raw Output
    The report consists of three major sections: (1) the header, which contains information about the query sequence, the database searched. (2) the one-line descriptions of each database sequence found to match the query sequence; these provide a quick overview for browsing; (3) the alignments for each database sequence matched (there may be more than one alignment for a database sequence it matches).
    In the one-line descriptions in the BLAST report, each line is composed of five fields: (a) UniProtKB accession number (b) UniProtKB ID, (c) the protein name. This line is often truncated in the one-line descriptions to keep the display compact; (d) the alignment score in bits. Higher scoring hits are found at the top of the list; and (e) the E-value, which provides an estimate of statistical significance. Adapted from The NCBI Handbook.

    A sample output for a BLAST search against the UniRef100 database is shown below



    1- Query Sequence on
    Click on this button to display the query sequence. Conversely, click it again to hide it.

    2- Save Results As
    Search results can be saved to the user's local computer. The results will be saved for selected entries or, if no proteins are selected, for all entries. Clicking "Table" will save the displayed columns as a tab-delimited text file, which may be imported into a spreadsheet for easier viewing or analysis. Clicking "FASTA" will save the IDs and sequences in FASTA format.

    3- BLAST, FASTA, Pattern Match, Multiple Alignment and Domain Display
    Retrieved entries can be further analyzed using the sequence analysis programs available in the Results page. First, select the protein(s) using the checkboxes on the left side of the table, then click the corresponding analysis tool.
  • Click "BLAST" or "FASTA" button, and a new query page will be displayed, along with the parameters that were selected in the initial search.
  • Click "Pattern Match" to search against the PROSITE database.
  • For multiple alignment, check at least 2 proteins (but no more than 70), then click the "Multiple Alignment" button. This will open the Multiple alignment form from which you can select one of the alignment programs: ClustalW, T-Coffee or Muscle. The result page will display the alignment and alignment viewer. For ClustalW and T-Coffee, the neighbor-joining tree and alignment can be viewed, edited and saved using either PIR-TAV viewer or JalView. For Muscle only Jalview is available.


  • 4- Results Display

    Sort Columns
    Columns can be sorted by the corresponding values by clicking on the arrow next to the column title. By default the table is sorted by Score.

    BLAST results are calculated for the representative member of each cluster, you may see the other members (but no BLAST results) by clicking on the "Show all members" logo. Results are shown in a table with the following default columns:

    UniRef100 ID
    It refers to the cluster identifier. Below this there is a link to the UniRef100 report with information on the UniRef100 cluster members, as well as links to the UniRef50 and UniRef90 clusters.

    Protein AC
    The Protein AC refers to the UniProtKB accession or UniParc ID. Below these identifiers, you may choose either the iProClass or the UniParc view of the protein report, respectively.

    Protein Name
    The common name given to a protein, that identifies its function or specifies its features.

    Length
    Number of amino acid residues in the peptide or protein.

    Organism Name
    The genus and species of the source organism from which the sequence originated. Links to NCBI taxonomy information are provided.

    BLAST Sequence Similarity Columns
    There are three columns related to BLAST results. The Alignment column, in the extreme right, shows the BLAST results in graphical format. The top bar represents the query sequence. The bars below show the region(s) on other sequences matched to the query sequence. The bar color indicates the magnitude of the BLAST score. Click on one of these bars to see its BLASTalignment paired with the query sequence.

    SSearch Columns
    SSearch is a pairwise implementation of the Smith-Waterman alignment algorithm. When two sequences are aligned, only the shared region is shown. Within the shared region, amino acid residues from one or both sequences can be aligned with either amino acids or gaps from the other sequence. The total length of the shared region, including gaps, is represented under the Overlap column. The percentage of identical residues in the alignment is given under %iden. Clicking on that number displays the SSearch full-length alignment.

    5- Show All Members
    By clicking this logo you will be able to see all sequences belonging to individual clusters.



    FASTA Search Help



    What is FASTA?
    FASTA can be used to search sequence databases, evaluate similarity scores, and identify periodic structures based on local sequence similarity. The FASTA program can compare a protein sequence to a DNA sequence database by translating the DNA database as it is searched. This search engine displays FASTA results (up to 200 matches), using the FASTA program (Pearson and Lipman, 1988) with default settings.

    Select a Database
    Perform FASTA search against:
  • UniProtKB is the central hub for the collection of functional information on proteins, with accurate, consistent, and rich annotation. It consists of two sections: a section containing manually-annotated records with information extracted from literature and curator-evaluated computational analysis (UniProtKB/Swiss-Prot), and a section with computationally analyzed records that await full manual annotation (UniProtKB/TrEMBL).
  • A subset of UniProtKB entries belonging to a certain organism or taxon group.
  • UniRef100 provides clustered sets of sequences at 100 % from UniProt Knowledgebase (including splice variants and isoforms) and selected UniParc records (mainly from PDB, Refseq and Ensembl databases), in order to obtain complete coverage of sequence space while hiding redundant sequences (but not their descriptions) from view. It combines identical sequences and sub-fragments with 11 or more residues (from any organism) into a single UniRef entry, displaying the sequence of a representative protein, the accession numbers of all the merged UniProtKB entries, and links to the corresponding UniProtKB and UniParc records.


  • FASTA Search Format
    The FASTA 'Search' input area accepts:

  • ">" followed by a UniProtKB sequence identifier. Spaces between letters in the input are not allowed, although spaces before or after the identifier are allowed.


  • Raw amino acid sequence format:
    MSEPQRLFFAIDLPAEIREQIIHWRATHFPPEAGRPVAADNLHLTLAFLGEVS
    AEKEKALSLLAGRIRQPGFTLTLDDAGQWLRSRVVWLGMRQPPRGLIQLAN
    MLRSQAARSGCFQSNRPFHPHITLLRDASEAVTIPPPGFNWSYAVTEFTLYA
    SSFARGRTRYTPLKRWALTQ

  • The query sequence intersperse with numbers and/or spaces, such as:
    1    msepqrlffa idlpaeireq iihwrathfp peagrpvaad nlhltlaflg evsaekekal
    61   sllagrirqp gftltlddag qwlrsrvvwl gmrqpprgli qlanmlrsqa arsgcfqsnr
    121  pfhphitllr daseavtipp pgfnwsyavt eftlyassfa rgrtrytplk rwaltq

  • Sequences are expected to be represented in the standard IUB/IUPAC amino acid codes, with these exceptions: lower-case letters are accepted and are mapped into upper-case; U and * are acceptable letters (see table) Numerical digits in the query sequence are automatically removed.

    FASTA Options
    -Expect
    The Expect (E-value) threshold establishes a statistical significance threshold for reporting database sequence matches. The default value is 0.0001, meaning that 0.0001 matches are expected to be found merely by chance. Lower Expect thresholds are more stringent, leading to fewer chance matches being reported. Increasing the expected threshold shows less stringent matches and is recommended when performing searches with short sequences as a short query is more likely to occur by chance in the database than a longer one, so even a perfect match (no gaps) can have low statistical significance and may not be reported. Increasing the Expect threshold allows you to look farther down in the hit list and see matches that would normally be discarded because of low statistical significance.

    References
      Pearson WR, and DJ Lipman (1988) Improved tools for biological sequence comparison. Proc. Natl. Acad. Sci. USA 85(8): 2444-2448.





    FASTA Results Help


    A sample output for a BLAST search against the UniProtKB database is shown below



    1- Query Sequence on
    Click on this button to display the query sequence. Conversely, click it again to hide it.

    2- Save Results As
    Search results can be saved to the user's local computer. The results will be saved for selected entries or, if no proteins are selected, for all entries. Clicking "Table" will save the displayed columns as a tab-delimited text file, which may be imported into a spreadsheet for easier viewing or analysis. Clicking "FASTA" will save the IDs and sequences in FASTA format.

    3- BLAST, FASTA, Pattern Match, Multiple Alignment and Domain Display
    Retrieved entries can be further analyzed using the sequence analysis programs available in the Results page. First, select the protein(s) using the checkboxes on the left side of the table, then click the corresponding analysis tool.
  • Click "BLAST" or "FASTA" button, and a new query page will be displayed, along with the parameters that were selected in the initial search.
  • Click "Pattern Match" to search against the PROSITE database.
  • For multiple alignment, check at least 2 proteins (but no more than 70), then click the "Multiple Alignment" button. This will open the Multiple alignment form from which you can select one of the alignment programs: ClustalW, T-Coffee or Muscle. The result page will display the alignment and alignment viewer. For ClustalW and T-Coffee, the neighbor-joining tree and alignment can be viewed, edited and saved using either PIR-TAV viewer or JalView. For Muscle only Jalview is available.
  • Domain display option, shows PFam domains (if present) in graphical format.


  • 4- Results Display

    Sort Columns
    Columns can be sorted by the corresponding values by clicking on the arrow next to the column title. By default the table is sorted by Score.

    The results from the search are displayed in a table with the following default columns:

    Protein AC/ID
    The Protein AC/ID refers to the UniProtKB accession number and ID, respectively. Below these identifiers, you may choose either the iProClass or the UniProtKB view of the protein report. The source of the UniProtKB sequence is shown as UniProtKB/Swiss-Prot or UniProtKB/TrEMBL if the protein sequence is from the Swiss-Prot or TrEMBL section of UniProtKB, respectively.

    Protein Name
    The common name given to a protein, that identifies its function or specifies its features.

    Length
    Number of amino acid residues in the peptide or protein.

    Organism Name
    The genus and species of the source organism from which the sequence originated. Links to NCBI taxonomy information are provided.

    PIRSF ID
    If a protein belongs to a PIRSF family, then this column will display the corresponding family identifier. Click on the ID to retrieve the PIRSF report (see annotated output).

    Sequence Similarity Columns
    There are three columns related to FASTA results. The Alignment column, in the extreme right, shows the FASTA results in graphical format. The top bar represents the query sequence. The bars below show the region(s) on other sequences matched to the query sequence. The bar color indicates the magnitude of the FASTA score. Click on one of these bars to see its FASTA alignment paired with the query sequence.



    Related Sequence Help

    Retrieve related sequences based on pre-computed BLAST results, therefore you will be able to have a glance at proteins similar to your query, significantly faster than running BLAST. This procedure is performed approximately every three months. Enter the UniProtKB sequence identifier and click "Search" button.

    Sample output for UniProtKB P53039



    1- E-value display
    Select the E-value cut-off to display your results.

    2- Query Sequence on
    Click on this button to display the query sequence. Conversely, click it again to hide it.

    3- Save Results As
    Search results can be saved to the user's local computer. The results will be saved for selected entries or, if no proteins are selected, for all entries. Clicking "Table" will save the displayed columns as a tab-delimited text file, which may be imported into a spreadsheet for easier viewing or analysis. Clicking "FASTA" will save the IDs and sequences in FASTA format.

    4- BLAST, FASTA, Pattern Match, Multiple Alignment and Domain Display
    Retrieved entries can be further analyzed using the sequence analysis programs available in the Results page. First, select the protein(s) using the checkboxes on the left side of the table, then click the corresponding analysis tool.
  • Click "BLAST" or "FASTA" button, and a new query page will be displayed, along with the parameters that were selected in the initial search.
  • Click "Pattern Match" to search against the PROSITE database.
  • For multiple alignment, check at least 2 proteins (but no more than 70), then click the "Multiple Alignment" button. This will open the Multiple alignment form from which you can select one of the alignment programs: ClustalW, T-Coffee or Muscle. The result page will display the alignment and alignment viewer. For ClustalW and T-Coffee, the neighbor-joining tree and alignment can be viewed, edited and saved using either PIR-TAV viewer or JalView. For Muscle only Jalview is available.
  • Domain display option, shows PFam domains (if present) in graphical format.


  • 5- Results Display

    Sort Columns
    Columns can be sorted by the corresponding values by clicking on the arrow next to the column title. By default the table is sorted by Score.

    The results from the search are displayed in a table with the following default columns:

    Protein AC/ID
    The Protein AC/ID refers to the UniProtKB accession number and ID, respectively. Below these identifiers, you may choose either the iProClass or the UniProtKB view of the protein report. The source of the UniProtKB sequence is shown as UniProtKB/Swiss-Prot or UniProtKB/TrEMBL if the protein sequence is from the Swiss-Prot or TrEMBL section of UniProtKB, respectively.

    Protein Name
    The common name given to a protein, that identifies its function or specifies its features.

    Length
    Number of amino acid residues in the peptide or protein.

    Organism Name
    The genus and species of the source organism from which the sequence originated. Links to NCBI taxonomy information are provided.

    PIRSF ID
    If a protein belongs to a PIRSF family, then this column will display the corresponding family identifier. Click on the ID to retrieve the PIRSF report (see annotated output).

    Sequence Similarity Columns
    There are three columns related to BLAST results. The Alignment column, in the extreme right, shows the BLAST results in graphical format. The top bar represents the query sequence. The bars below show the region(s) on other sequences matched to the query sequence. The bar color indicates the magnitude of the BLAST score. Click on one of these bars to see its alignment paired with the query sequence.



    Peptide Match


    Find an exact match for a peptide sequence query in the selected database. You can perform the search against:
  • UniProtKB is the central hub for the collection of functional information on proteins, with accurate, consistent, and rich annotation. It consists of two sections: a section containing manually-annotated records with information extracted from literature and curator-evaluated computational analysis (UniProtKB/Swiss-Prot), and a section with computationally analyzed records that await full manual annotation (UniProtKB/TrEMBL).
  • A subset of UniProtKB entries belonging to a certain organism or a set of organisms.



  • Peptide Match Result page



    1- Query Peptide
    Show the query peptide and allow user to do a new search by submitting a new query peptide.

    2- Save Results As
    The output can be saved to the user's local computer. The results will be saved for selected entries or, if no proteins are selected, for all entries. Clicking "Table" will save the displayed columns as a tab-delimited text file, which may be imported into a spreadsheet for easier viewing or analysis. Clicking "FASTA" will save the IDs and sequences in FASTA format.

    3- Analyze: BLAST, FASTA, Pattern Match, Multiple Alignment and Domain Display
    Retrieved entries can be further analyzed using the sequence analysis programs available in the Results page. First, select the protein(s) using the checkboxes on the left side of the table, then click the corresponding analysis tool.
  • Click "BLAST" or "FASTA" button, and a new query page will be displayed, along with the parameters that were selected in the initial search.
  • Click "Pattern Match" to search against the PROSITE database.
  • For multiple alignment, check at least 2 proteins (but no more than 70), then click the "Multiple Alignment" button. This will open the Multiple alignment form from which you can select one of the alignment programs: ClustalW, T-Coffee or Muscle. The result page will display the alignment and alignment viewer. For ClustalW and T-Coffee, the neighbor-joining tree and alignment can be viewed, edited and saved using either PIR-TAV viewer or JalView. For Muscle only Jalview is available.
  • Domain display option, shows PFam domains (if present) in graphical format.

  • 4- Results Display
    Results of the search are displayed in a table.

    Protein AC/ID
    The Protein AC/ID refers to the UniProtKB or UniRef100 identifiers. Below these numbers, you may choose either the iProClass or the UniProtKB/UniRef100 view of each protein report. The source of the UniProtKB sequence is shown as UniProtKB/Swiss-Prot or UniProtKB/TrEMBL if the protein sequence is from Swiss-Prot or TrEMBL, respectively.

    Protein Name
    The common name given to a protein, that identifies its function or specifies its features.

    Length
    Number of amino acid residues in the peptide or protein.

    Organism Name
    The genus and species of the source organism from which the sequence originated. Links to NCBI taxonomy information are provided.

    PIRSF ID
    If a protein belongs to a PIRSF family, then this column will display the corresponding family identifier. Click on the ID to retrieve the PIRSF report (see annotated output).

    Match Range
    This column displays in red the peptide query within the sequence.

    5- Multiple organisms
    Click to show the taxonomy IDs of matched organisms.

    6- Browse by taxonomy group
    Click to show the taxonomy groups of matched sequences as shown below.


    Batch Peptide Match

    Find the exact match for a set of query peptide sequences in the selected database: You can perform the search against:
  • UniProtKB is the central hub for the collection of functional information on proteins, with accurate, consistent, and rich annotation. It consists of two sections: a section containing manually-annotated records with information extracted from literature and curator-evaluated computational analysis (UniProtKB/Swiss-Prot), and a section with computationally analyzed records that await full manual annotation (UniProtKB/TrEMBL).
  • A subset of UniProtKB entries belonging to a certain organism or a set of organisms.

  • The batch peptide match results can be retrieved as a text file for further analysis. The match results are organised based on the query peptide sequences.




    Pattern Search


    A pattern is a formula (regular expression) that represents the conserved region of a group of related proteins. Once constructed, the pattern is used by a pattern-matching program to find possible occurrences of the conserved region in the sequence database.

    PROSITE is a database that contains patterns and profiles specific for more than a thousand protein families or domains. Each of these signatures comes with documentation providing background information on the structure and function of these proteins.

    Identification of patterns in a protein or group of proteins can help in assessing protein function or predict a certain post-translational modification. However, pattern search results should be further investigated, since due to the nature of the approach the patterns are often too specific (many false negatives) or insufficiently selective (high probability of occurrence). As is the case, for example, for the PROSITE pattern:
    PS00001
    ID ASN_GLYCOSYLATION; PATTERN.
    DE N-glycosylation site.
    PA N-{P}-[ST]-{P}
    This pattern can be found in most proteins, however, N-glycosylation is a modification that takes place in the Endoplasmic Reticulum during synthesis of membrane and secreted proteins.

    Pattern search at PIR allows:
  • The search of PROSITE patterns (note that we only include patterns, profiles are excluded) in a query sequence, entering the single amino acid code sequence or its unique ID
  • The search of a specific PROSITE or user-defined pattern against one of the following sequence database:
    (i)UniProtKB is the central hub for the collection of functional information on proteins, with accurate, consistent, and rich annotation. It consists of two sections: a section containing manually-annotated records with information extracted from literature and curator-evaluated computational analysis (UniProtKB/Swiss-Prot), and a section with computationally analyzed records that await full manual annotation (UniProtKB/TrEMBL).
    (ii) A subset of UniProtKB entries belonging to a certain organism or taxon group.
    (iii) UniRef100 provides clustered sets of sequences at 100 % from UniProt Knowledgebase (including splice variants and isoforms) and selected UniParc records (mainly from PDB, Refseq and Ensembl databases), in order to obtain complete coverage of sequence space while hiding redundant sequences (but not their descriptions) from view. It combines identical sequences and sub-fragments with 11 or more residues (from any organism) into a single UniRef entry, displaying the sequence of a representative protein, the accession numbers of all the merged UniProtKB entries, and links to the corresponding UniProtKB and UniParc records.


  • How to write a protein pattern
    Follow the example below to build the protein pattern.
    [LIVM]-[VIC]-x(2) -G-[DENQTA]-x-[GAC]-x(2) -[LIVMFY](4)-x(2)-G
    1. Use capital letters for amino acid residues and put a "-" between two amino acids (not required).

    2. Use "[] for a choice of multiple amino acids in a particular position.
      • [LIVM] means that L, I, V, or M can be in the first position.

    3. Use "{…}" to exclude amino acids.
      • {CF} means C and F should not be in that particular position

    4. Use "x" for a position that can be any amino acid.

    5. Use "(n)", where n is a number, for multiple positions.
      • x (3) is the same as "xxx"

    6. Use "(n1,n2)" for multiple or variable positions.
      • " x (1,4) represents "x" or "xx" or "xxx" or "xxxx"

    7. Use the symbol ">" at the beginning or end of the pattern to require the pattern to match the N or C terminus.
      • ">MDEL" finds only sequences that start with MDEL
      • "DEL>" finds only sequences that end with DEL
    [LIVM]-[VIC]-x(2) -G-[DENQTA]-x-[GAC]-x(2) -[LIVMFY](4)-x(2)-G
    Illustrates a 17 amino acid peptide that has: an L, I, V, or M at position 1; a V, I, or C at position 2; any residue at positions 3 and 4; a G at position 5 and so on ….

    Note: You can also write the above pattern as:
    [LIVM] [VIC] x (2) G [DENQTA] x [GAC] x (2) [LIVMFY] (4) x (2) G

    Result for proteins with PROSITE pattern PS00888 (Cyclic nucleotide-binding domain signature 1)



    1- Save Results As
    The output can be saved to the user's local computer. The results will be saved for selected entries or, if no proteins are selected, for all entries. Clicking "Table" will save the displayed columns as a tab-delimited text file, which may be imported into a spreadsheet for easier viewing or analysis. Clicking "FASTA" will save the IDs and sequences in FASTA format.

    2- BLAST, FASTA, Pattern Match, Multiple Alignment and Domain Display
    Retrieved entries can be further analyzed using the sequence analysis programs available in the Results page. First, select the protein(s) using the checkboxes on the left side of the table, then click the corresponding analysis tool.
  • Click "BLAST" or "FASTA" button, and a new query page will be displayed, along with the parameters that were selected in the initial search.
  • Click "Pattern Match" to search against the PROSITE database.
  • For multiple alignment, check at least 2 proteins (but no more than 70), then click the "Multiple Alignment" button. This will open the Multiple alignment form from which you can select one of the alignment programs: ClustalW, T-Coffee or Muscle. The result page will display the alignment and alignment viewer. For ClustalW and T-Coffee, the neighbor-joining tree and alignment can be viewed, edited and saved using either PIR-TAV viewer or JalView. For Muscle only Jalview is available.
  • Domain display option, shows PFam domains (if present) in graphical format.


  • 3- Results Display
    Results of the search are displayed in a table. Columns can be sorted by the corresponding values by clicking on the arrow next to the column title.

    Protein AC/ID
    The Protein AC/ID refers to the UniProtKB accesion and identifier, respectively. Below these, you may choose either the iProClass or the UniProtKB view of each protein report. The source of the UniProtKB sequence is shown as UniProtKB/Swiss-Prot or UniProtKB/TrEMBL if the protein sequence is from Swiss-Prot or TrEMBL, respectively.

    Protein Name
    The common name given to a protein, that identifies its function or specifies its features.

    Length
    Number of amino acid residues in the peptide or protein.

    Organism Name
    The genus and species of the source organism from which the sequence originated. Links to NCBI taxonomy information are provided.

    PIRSF ID
    If a protein belongs to a PIRSF family, then this column will display the corresponding family identifier. Click on the ID to retrieve the PIRSF report (see annotated output).

    Match Range
    This column displays the sequence range for the query pattern.

    Partial result for PROSITE patterns in UniProtKB O05689






    Multiple Alignment Help





    Select the alignment program: ClustalW, T-Cofee or Muscle. Then, enter multiple sequences (no more than 70) with their corresponding ID lines in FASTA format, or enter multiple UniProtKB IDs separated by lines, commas or spaces. Then click the "Submit" button.
    The result page will display the alignment and alignment viewer. For ClustalW and T-Coffee, the neighbor-joining tree and alignment can be viewed, edited and saved using either PIR-TAV viewer or JalView (see below). For Muscle only Jalview is available.



    See sample output

    FASTA Format
    A sequence in FASTA format begins with a single-line description, followed by lines of sequence data. The description line is distinguished from the sequence data by a greater-than (">") symbol in the first column. One benefit of using FASTA format is that the sequence identifier will be reported with the results. An example sequence in FASTA format is:

    >gi|3287971|sp|P37025|LIGT_ECOLI 2'-5' RNA ligase MSEPQRLFFAIDLPAEIREQIIHWRATHFPPEAGRPVAADNLHLTLAFLGEVSAEK EKALSLLAGRIRQPGFTLTLDDAGQWLRSRVVWLGMRQPPRGLIQLANMLRSQA ARSGCFQSNRPFHPHITLLRDASEAVTIPPPGFNWSYAVTEFTLYASSFARGRTRY TPLKRWALTQ




    Pairwise Alignment Help



    Insert two sequences using the single letter amino acid code or enter two UniProtKB identifiers codes. Press the "Submit" button and the alignment results will show the SSEARCH Smith-Waterman full-length alignments between two sequences (SSEARCH program version 3.4t24 July 21, 2004).

    References
      Smith TF, and MS Waterman (1981) Identification of common molecular subsequences. J. Mol. Biol. 147, 195-197.


    Pairwise alignment of P53039 and Q6FQ69




    ID Mapping Help


    This function allows the mapping of IDs between UniProtKB and other databases

    First specify the type of ID of your source data (multiple types of ID can be selected by using the ctrl key). Specify the type of ID you want the mapping to. Type or paste the identifiers in the box provided separated by a space or return key. Alternatively, you can upload a file with a list of IDs. Select the output format and finally, press "Map".

    The result page contains the list of ID matches. The IDs can be cut and pasted if needed or saved as a text file using the "save as" option provided by your web browser.
    See annotated output below.

    Mapping of GI numbers to UniProt ACs






    Composition/Molecular Weight Calculation Help



    Retrieve the composition information and the molecular weight of a single or multiple sequences based on the UniProtKB identifier or the single letter amino acid code. The results contain the number of residues and percentage for each amino acid in the sequence. The calculation notes for the molecular weight and the molecular weight for each amino acid are also given.

    Molecular weight composition for P53039




    PIRSF Scan


    Search your query sequence against the fully curated PIRSF families. When you submit your query sequence, it will be searched against the full-length and domain HMM models for the fully curated PIRSFs by HMMER program. If a match is found, the matched regions and statistics will be displayed. The query sequence should be entered using single letter code or a UniProtKB identifier.

    PIRSF scan for Q8Y5X7






    Amino Acid Sequence Code Table



    Code Amino Acid
    Aalanine
    Baspartate or asparagine
    Ccysteine
    Daspartate
    Eglutamate
    Fphenylalanine
    Gglycine
    Hhistidine
    Iisoleucine
    Klysine
    Lleucine
    Mmethionine
    Nasparagine
    Opyrrolysine
    Pproline
    Qglutamine
    Rarginine
    Sserine
    Tthreonine
    Uselenocysteine
    Vvaline
    Wtryptophan
    Ytyrosine
    Zglutamic acid or glutamine
    BLAST and FASTA also accept
    Xany
    *translation stop







    UniProtKB Identifiers for BLAST/FASTA Search and Analysis Tools


    The following table shows examples for UniProtKB accession number and ID

    Field Example
    UniProtKB ID IMDH2_HUMAN
    UniProtKB Accession P12268




    iProClass Report

    PIRSF Report
    Sample report for PIRSF000186. Information about the numbered fields is provided below.
    1- PIRSF Hierarchy DAG View
    2- PIRSF Taxonomic Distribution
    3- Multiple Alignment
    4- Domain Architecture

    Master Protein Directory Help


    This section describes the Master Protein Directory Text Search page. The Master Directory contains information on proteins and reagents identified by NIAID’s Biodefense Proteomics Research Centers. A description of this research program can be found here. The overall layout of the page is similar to the regular PIR text search results however; it differs in the default columns displayed and has additional attributes to search. There are two columnar displays; 1) default display which is used to display multiple data types together (i.e. MS_proteomic and Microarray); and, 2) data type specific which is used when only one data type is selected and contains additional information that applies to that data type only. To view the data specific display from the pull-down menu --Select A Data Type to Show-- select a data type.

    Default Display for Multiple Data Types



    Data Type Specific Display (Microarray)



    Search and Results Options


    1- Browse Selection
    By default the directory shows all data. This feature, a pull-down menu of options at the top of the page, allows you restrict your browsing by 1) data type, 2) Proteomics Research Center and 3) Organism

    2- Search
    Allows you to perform an additional search in case you want to further filter your output or you want to start a new search (no need to go back to the previous page). The available options are shown in a pull-down menu and are the same protein attributes as those found in the iProClass database with the addition of directory specific attributes listed under the – Master Directory – heading. Enter any text string or substring, case does not matter. If you wish to determine just the presence or absence of a value use the words “not null” or “null”. Wildcards are allowed.

    3- Display Options
    Depending on your specific need you can choose the columns to be displayed. To do this, click on the "Display Option" button, select the relevant field(s) in the "Fields Not in Display" list and transfer them to the "Fields in Display" list via the ">" button. Conversely, columns can be removed from display. Finally, click on "apply" for the changes to take effect.

    4- Save Results As
    The output can be saved to the user's local computer. The results will be saved for selected entries or, if no proteins are selected, for all entries. Clicking "Table" will save the displayed columns as a tab-delimited text file, which may be imported into a spreadsheet for easier viewing or analysis. Clicking "FASTA" will save the IDs and sequences in FASTA format.


    Data Type Specific Display (Microarray)



    5- Analyze: BLAST, FASTA, Pattern Match, Multiple Alignment and Domain Display
    Retrieved entries can be further analyzed using the sequence analysis programs available in the Results page. First, select the protein(s) using the checkboxes on the left side of the table, then click the corresponding analysis tool.
  • Click "BLAST" or "FASTA" button, and a new query page will be displayed, along with the parameters that were selected in the initial search.
  • Click "Pattern Match" to search against the PROSITE database.
  • For multiple alignment, check at least 2 proteins (but no more than 50), then click the "Multiple Alignment" button. A ClustalW alignment and neighbor-joining tree will be generated along with a link to the interactive PIR viewer.
  • Domain display option, shows PFam domains (if present) in graphical format.

    6- Results Display
    Results of the search are displayed in a customizable table. The exact columns displayed will depend on the fields searched for, and user preference.

    Protein AC/ID
    The Protein AC/ID refers to the UniProtKB or UniParc identifiers. Below these numbers, you may choose either the iProClass or the UniProtKB/UniParc view of the protein report. The source of the UniProtKB sequence is shown as UniProtKB/Swiss-Prot or UniProtKB/TrEMBL if the protein sequence is from Swiss-Prot or TrEMBL section of UniProt Knowledgebase, respectively., respectively. Alternatively, the UniParc ID will be displayed if the sequence is not present in the UniProtKB database but is present in UniParc.

    Dir.ID
    The Directory ID is a unique identifier selected from the data submitted by the Proteomic Research Center. It is used in the master directory to identify a specific research result and to link protein information in the Master Directory to the detailed results in the Protein Bank.

    Center
    Identifies the NIAID Biodefense Proteomics Research Center that produced the data. Clicking the name show additional information on the center and data they submitted.

    Data Type
    Identifies the type of data provided. Clicking the Data type name opens a new window with protein information shown in the data type specific format.

    Experiment #
    Links to related experimental data in the protein data bank.

    Protein Name
    The common name given to a protein, that identifies its function or specifies its features.

    Length
    Number of amino acid residues in the peptide or protein.

    Organism Name
    The genus and species of the source organism from which the sequence originated. Links to NCBI taxonomy information are provided.

    PIRSF ID
    If a protein belongs to a PIRSF family, then this column will display the corresponding family identifier. Click on the ID to retrieve the PIRSF report (see annotated output).

    7– Data Type Specific Search Options
    All the common fields listed above can be searched as can any optional columns a user might add. In addition there are data type specific fields that are only displayed in the datatype specific display but can also be searched. Note: some reagent specific fields may be empty.

    Searchable fields are:
  • AA Range – Used when data pertains to a specific domain only. Currently used for structures.
  • Comment – Project and protein specific annotation.
  • IEDB ID – Immune Eptitope ID, links to the Immune Epitope Database & Analysis Resource
  • Expression Condition – A controlled vocabulary currently used for Microarray and Mass Spectrometry Data. Selecting this option will provide a list of allowable terms. Though terms are shared between experiments not all terms exist in all experiments.
  • Expression Status – A controlled vocabulary currently used for Microarray and Mass Spectrometry Data. Selecting this option will provide a list of allowable terms. Though terms are shared between experiments not all terms exist in all experiments.
  • GO Slim Func. – GO slim biological functional terms associated with the protein.
  • GO Slim Comp - GO slim biological compartment terms associated with the protein.
  • GO Slim Proc. - GO slim biological process terms associated with the protein.
  • Insertion Point – Used for clones to indicate the insertion site on the vector.
  • Interaction ID – Unique ID for interaction.
  • Clone ID – Unique ID for the clones insert.
  • PDB ID – Protein Data Bank ID for protein structures.
  • Prey AC – iProclass Accession for prey protein in a Y2H interacting pair
  • Publication – PRC publication related to the protein.
  • Vector – Name of cloning Vector
  • BRC Link – Bioinformatics Resource Center link. Protein specific link to one of the NIAID funded BRC centers database.

    Complete Predicted Proteomes Help


    This tool allows text searches restricted to complete proteomes as annotated by the UniProt Consortium and being studied by the Biodefense Proteomics Research Program. The results are linked to the Master Protein Directory which contains experimental information about some of the proteins.

    To begin select the proteomes of interest and search.



    Search and Results Options


    1- Search
    Once in the tool you can further restrict your search. The available options are shown in a pull-down menu and are the same protein attributes as those found in the iProClass database with the addition of the - Master Directory – (MD) option . Enter any text string or substring, case does not matter. If you wish to determine just the presence (+) or absence (blank) of a value use the words “not null” or “null”. Wildcard searches are allowed.

    2- Display Options
    Depending on your specific need you can choose the columns to be displayed. To do this, click on the "Display Option" button, select the relevant field(s) in the "Fields Not in Display" list and transfer them to the "Fields in Display" list via the ">" button. Conversely, columns can be removed from display. Finally, click on "apply" for the changes to take effect.

    3- Save Results As
    The output can be saved to the user's local computer. The results will be saved for selected entries or, if no proteins are selected, for all entries. Clicking "Table" will save the displayed columns as a tab-delimited text file, which may be imported into a spreadsheet for easier viewing or analysis. Clicking "FASTA" will save the IDs and sequences in FASTA format.



    4- Analyze: Master Directory, BLAST, FASTA, Pattern Match, Multiple Alignment and Domain Display
    Retrieved entries can be further analyzed using the Master Directory or using the sequence analysis programs available in on the PIR website. Master Directory: Any protein with information present in the Master Directory of the Biodefense Proteomics Research Program is indicated with a (+) in the MD column. Checking these entries and then selecting ‘Master Directory’ in the analyze tool bar opens the Master Directory with the selected proteins shown. Sequence Analysis: Selecting these options sends you to the PIR website where the analysis is performed. To return to the Master Directory use your browser back button. First, select the protein(s) using the checkboxes on the left side of the table, then click the corresponding analysis tool.
  • Click "BLAST" or "FASTA" button, and a new query page will be displayed, along with the parameters that were selected in the initial search.
  • Click "Pattern Match" to search against the PROSITE database.
  • For multiple alignment, check at least 2 proteins (but no more than 50), then click the "Multiple Alignment" button. A ClustalW alignment and neighbor-joining tree will be generated along with a link to the interactive PIR viewer.
  • Domain display option, shows PFam domains (if present) in graphical format.

    5- Results Display
    Results of the search are displayed in a customizable table. The exact columns displayed will depend on the fields searched for, and user preference.

    Protein AC/ID
    The Protein AC/ID refers to the UniProtKB or UniParc identifiers. Below these numbers, you may choose either the iProClass or the UniProtKB/UniParc view of the protein report. The source of the UniProtKB sequence is shown as UniProtKB/Swiss-Prot or UniProtKB/TrEMBL if the protein sequence is from Swiss-Prot or TrEMBL section, respectively. Alternatively, the UniParc ID will be displayed if the sequence is no present in the UniProtKB database along with the UniParc report.

    MD (Master Directory)
    Any protein with information present in the Master Directory of the Biodefense Proteomics Research Program is indicated with a (+) in the MD column. To restrict your search to only proteins that are present in the MD select “Master Directory” in the search field options and enter the value “yes” or “not null”.

    Protein Name
    The common name given to a protein, that identifies its function or specifies its features.

    Length
    Number of amino acid residues in the peptide or protein.

    Organism Name
    The genus and species of the source organism from which the sequence originated. Links to NCBI taxonomy information are provided.

    PIRSF ID
    If a protein belongs to a PIRSF family, then this column will display the corresponding family identifier. Click on the ID to retrieve the PIRSF report (see annotated output).

    Related Seq.
    This column shows the number of pre-computed BLAST hits obtained using default parameters. Only up to 300 sequences will be displayed. By clicking the number you access to the related sequence page. This allows to have a glance at sequence similarity in a very fast way. The number itself already provides some information about how unique the protein is. For example, a very low number may tell you that the query is specific to a certain species, genus, taxon, etc. The "+" sign next to the Related Sequence title allows to compare number of related sequences at 3 different E-value cut-offs as shown below.




    GO Slim Analysis

    GO slims are smaller versions of the Gene Ontologies containing a subset of the terms in the whole GO. They give a broad overview of the ontology content without the detail of the specific fine grained terms. GO slims are particularly useful for giving a summary of the results of GO annotation of a genome or proteome when broad classification of gene product function is required.

    You can view the GO slim terms for Biological Function, Component, and Process by selecting the “Show GO Slim” button in the analysis tool bar. You can then view statistics for the individual ontologies (Function, Component, and Function) by checking entries of interest and selecting the ontology to show. Follow the GO ID links to learn more about the GO term.




    Master Reagent Directory Help


    This section describes the Master Reagent Directory Text Search page. The Reagent Directory contains reagents identified by NIAID’s Biodefense Proteomics Research Centers and provides links to repositories were reagents can be obtained. A description of the proteomics research program can be found here. The overall layout of the page is similar to the regular PIR text search results however; it differs in the default columns displayed and has additional attributes to search. The are two columnar displays; 1) default display which is used to display multiple reagent types together (i.e. antibodies and clones); and, 2) reagent type specific which is used when only one reagent type is selected and contains additional information that applies to that reagent only.

    Default Display for Multiple Reagent Types



    Reagent Type Specific Display (Antibody)



    Search and Results Options


    1- Browse Selection
    By default the directory shows all reagents. This feature, a pull-down menu of options, allows you restrict your browsing by 1) Reagent type, 2) Proteomics Research Center and 3) Organism

    2- Search
    Allows you to perform an additional search in case you want to further filter your output or you want to start a new search (no need to go back to the previous page). The available options are shown in a pull-down menu (see current options below). Enter any text string or substring, case does not matter. If you wish to determine just the presence or absence of a value use the words “not null” or “null”.

    3- Display Options
    Depending on your specific need you can choose the columns to be displayed. To do this, click on the "Display Option" button, select the relevant field(s) in the "Fields Not in Display" list and transfer them to the "Fields in Display" list via the ">" button. Conversely, columns can be removed from display. Finally, click on "apply" for the changes to take effect.

    4- Save Results As
    The output can be saved to the user's local computer. The results will be saved for selected entries or, if no reagents are selected, for all entries. Clicking "Table" will save the displayed columns as a tab-delimited text file, which may be imported into a spreadsheet for easier viewing or analysis.



    5- Results Display
    Results of the search are displayed in a customizable table. The exact columns displayed will depend on the fields searched for, and user preference. The default columns are described below.

    Reagent ID
    The Reagent ID refers is a unique identifier. If the Reagent is deposited in the BEI resource is the same ID used at BEI.

    Reagent Type
    Identifies the type of reagent. Clicking the reagent type name opens a new window with reagent information shown in the reagent specific format.

    Name/Description
    Descriptive text about the reagent.

    Organism Name
    The genus and species name of the source organism from which or for which the reagent was developed. For example: Bacterial strains list the bacterial species name here, not the strain. Antibodies though originating from mouse or rabbit list the antigens organism. Links to NCBI taxonomy information are provided.

    Center
    Identifies the NIAID Biodefense Proteomics Research Center that produced the reagent. Clicking the name show additional information on the center and data they submitted.

    Experiment #
    Links to related experimental data in the protein data bank. If these reagents were used for a particular set of experiments this number would link to that data.

    Publication
    Links to PRC publications that used the reagent.

    Source
    Links to reagent repositories where users can obtain the reagent.

    6- Reagent Specific Search Options
    All the common fields listed above can be searched. In addition there are reagent specific fields that are only displayed in the reagent specific display but can also be searched. Note: some reagent specific fields may be empty.

    Searchable fields are:
  • Taxonomy ID – NCBI Taxonomy ID.
  • Ab EBPC – Internal tracking number for Einstein Biodefense Proteomics Center
  • Ab ELISA Data – Elisa titers
  • Ab EM? – Value = Y, N, null. Y means spot detected in Electron Microscopy.
  • Ab IFA? - Value = Y, N, null. Y means spot detected in Immunofluorescence Assay.
  • Ab Type – Value = Monoclonal, Polyclonal, null
  • Ab Western? - Y, N, null. Y means band detected in Western blot.
  • Ab Peptide – Peptides used to immunize and for Eliza
  • Bac Parent ID – Parent Strain Identifier
  • Bac Strain ID – Reagent Strain ID
  • Clone Insertion Point – Insertion point on clone vector.
  • Clone Locus ID – Locus ID of insert sequence.
  • Clone Vector ID – Name of Vector used
  • Clone Protein ID – iProclass accession or protein, links to Master Protein Directory.


    Representative Proteomes Help

    1. What are Representative Proteomes?
    Representative Proteomes (RPs), are proteomes that are selected from Representative Proteome Groups (RPGs) containing similar proteomes calculated based on co-membership in UniRef50 clusters. Representative Proteome is the proteome that can best represent all the proteomes in its group in terms of the majority of the sequence space and information. RPs at 75%, 55%, 35% and 15% co-membership threshold are provided to allow users to decrease or increase the granularity of the sequence space based on their requirements.

    2. Representative Proteomes BLAST search.
    1. Select a Database BLAST search can be performed against RP75, RP55, RP35 or RP15: Lower RPs (such as RP15) has fewer proteomes than higher RPs (such as RP75). For example, to get the least number of BLAST results choose RP15. For additional help on BLAST please see http://pir.georgetown.edu/pirwww/support/help.shtml#3.

    3. Availability and usage of RP.
    For the threshold values of 75, 55, 35, and 15 (RP75, RP55, RP35 and RP15), corresponding Representative Proteome Group files are provided, via links from the RP home page, in the format as below:

    >rp_taxon_id  rp_code  rp_name  rp_annotation_score (AS)  taxon_group C(THRESHOLD)
     mp_taxon_id  mp_code  mp_name  mp_annotation_score (AS)  taxon_group X_to_rp(X)
       ...
    >rp_taxon_id  rp_code  rp_name  rp_annotation_score (AS)  taxon_group C(THRESHOLD)
       ...
    Where 
    rp is Representative Proteome,
    mp is Member Proteome in the Representative Proteome Group
    
    An example of Representative Proteome Group is shown below:
    >205920  EHRCR  Ehrlichia chaffeensis (strain Arkansas)     Bac/Alpha-proteo  1111.19332(AS)  55(CUTOFF)
     269484  EHRCJ  Ehrlichia canis (strain Jake)               Bac/Alpha-proteo  1111.10824(AS)  71.20366(X)
     302409  EHRRG  Ehrlichia ruminantium (strain Gardel)       Bac/Alpha-proteo  1111.10730(AS)  64.05622(X)
     254945  EHRRW  Ehrlichia ruminantium (strain Welgevonden)  Bac/Alpha-proteo  1111.12521(AS)  65.56531(X)
    
    
    Also provided are the sequence files in FASTA format for the RP75, RP55, RP35 and RP15 sets. Users can choose to make their own customized RP set by using the taxon-based table or perl script available via a link from the RP home page. For example, we suspect that, for some users, the ideal set could be RP75 for Animals + RP55 for other cellular organisms + any missing GO Reference Genomes.

    4. Representative Proteome availability from the iProClass interface.
    iProClass is an integrated data-warehouse containing all UniProtKB proteins and additional proteins from NCBI resources. The proteins from the defined representative proteome sets are indexed in iProClass and are available for BLAST (http://proteininformationresource.org/rps/blast_rp.shtml). Additionally, all the proteins from the RP55 set can be retrieved from http://proteininformationresource.org/pirwww/search/textsearch.shtml by selecting Rep Proteome and then typing in not null and clicking on Search. Users can perform additional filtering on the retrieved set by performing Boolean searches using more than 65 fields available from the text search pull down menu. The BLAST and the text search results can be downloaded from the results page for further analysis.

    5. Browsing the Representative Proteomes.
    The RPs at the four different thresholds can be viewed at http://proteininformationresource.org/cgi-bin/rps_tree.pl. The top most nodes are Archaea, Bacteria and Eukaryota and the fully expanded view shows all the proteomes that have been analyzed to identify the RPs. Browsing the RPs at different threshold for different taxonomy nodes can provide clues as to which CMT is best for a particular branch and how the RPs are distributed in the taxonomy tree. Once a desired set of RPs is displayed on the screen, it can be printed for future reference.

    6. Representative Proteomes data update.
    The protein sets used for text search and BLAST search are updated every four weeks. New proteomes are added every six months. All releases are archived for at least 5 years.



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