The UniProt knowledgebase

www..org

a hub of integrated data

[email protected] Swiss-Prot group, Geneva SIB Swiss Institute of

Science cover, february 2011 data knowledge protein sequence functional information UniProt consortium

EBI : European Bioinformatics Institute (UK) SIB : Swiss Institute of Bioinformatics (CH) PIR : Protein information resource (US) www.uniprot.org UniProt databases UniProtKB: protein sequence knowledgebase, 2 sections UniProtKB/Swiss-Prot and UniProtKB/TrEMBL (query, Blast, download) (~15 mo entries)

UniParc: protein sequence archive (ENA equivalent at the protein level). Each entry contains a protein sequence with cross- links to other databases where you find the sequence (active or not). Not annotated (query, Blast, download) (~25 mo entries)

UniRef: 3 clusters of protein sequences with 100, 90 and 50 % identity; useful to speed up sequence similarity search (BLAST) (query, Blast, download) (UniRef100 10 mo entries; UniRef90 7 mo entries; UniRef50 3.3 mo entries)

UniMES: protein sequences derived from metagenomic projects (mostly Global Ocean Sampling (GOS)) (download) (8 mo entries, included in UniParc) UniProt databases The central piece UniProtKB an encyclopedia on

composed of 2 sections UniProtKB/TrEMBL and UniProtKB/Swiss-Prot unreviewed and reviewed automatically annotated and manually annotated

released every 4 weeks

UniProtKB Origin of protein sequences

UniProtKB protein sequences are mainly derived from

- INSDC (translated submitted coding sequences - CDS) 85 % - Ensembl (gene prediction ) and RefSeq sequences - Sequences of PDB structures 15 % - Direct submission or sequences scanned from literature

Notes: - UniProt is not doing any gene prediction - Most non-germline immunoglobulins, T-cell receptors , most patent sequences, highly over-represented data (e.g. viral antigens), pseudogenes sequences are excluded from UniProtKB, - but stored in UniParc - Data from the PIR database have been integrated in UniProtKB since 2003.

EMBL Manual annotation of Swiss-Prot the sequence and associated biological information TrEMBL

Automated extraction of protein sequence (translated CDS), gene name and references. Automated annotation UniProtKB/TrEMBL

unreviewed Automatic annotation

released every 4 weeks

Protein and gene names Taxonomic information Automated annotation Function, Subcellular location, Catalytic activity, Sequence similarities…

References Automated annotation One protein sequence transmembrane domains, One species signal peptide…

Cross-references Automated annotation to over 125 databases UniProtKB/TrEMBL Keywords and www.uniprot.org

UniProtKB/TrEMBL

Automatic annotation Protein sequence - The quality of the protein sequences is dependent on the information provided by the submitter of the original nucleotide entry (CDS) or of the gene prediction pipeline (i.e. Ensembl). - 100% identical sequences (same length, same organism are merged automatically).

Biological information Sources of annotation - Provided by the submitter (EMBL, PDB, TAIR…) - From automated annotation (automated generated annotation rules (i.e. SAAS) and/or manually generated annotation rules (i.e. UniRule))

UniProtKB/TrEMBL

Example of fully automatic annotation: SAAS

• Rules are derived from the UniProtKB/Swiss-Prot manual annotation.

• Fully automated rule generation based on C4.5 decision tree algorithm.

• One annotation, one rule.

• High stringency – require 99% or greater estimated precision to generate annotation (test on UniProtKB/Swiss-Prot)

• Rules are produced, updated and validated at each release.

UniProtKB/Swiss-Prot

reviewed manually annotated

released every 4 weeks

Manual annotation Function, Subcellular location, Protein and gene names Catalytic activity, Disease, Taxonomic information Tissue specificty, Pathway…

MSKEKFERTKPHVNVGTIGHVDHGKTTLTAAITTVLAKTYGGAAR AFDQIDNAPEEKARGITINTSHVEYDTPTRHYAHVDCPGHADYVK References NMITGAAQMDGAILVVAATDGPMPQTREHILLGRQVGVPYIIVFLOne protein sequence Manual annotation NKCDMVDDEELLELVEMEVRELLSQYDFPGDDTPIVRGSALKALE Post-translational modifications, GDAEWEAKILELAGFLDSYIPEPERAIDKPFLLPIEDVFSISGRGOne gene TVVTGRVERGIIKVGEEVEIVGIKETQKSTCTGVEMFRKLLDEGROne species variants, transmembrane domains, AGENVGVLLRGIKREEIERGQVLAKPGTIKPHTKFESEVYILSKD signal peptide… EGGRHTPFFKGYRPQFYFRTTDVTGTIELPEGVEMVMPGDNIKMV VTLIHPIAMDDGLRFAIREGGRTVGAGVVAKVLG

Alternative products: protein sequences produced by alternative splicing, alternative promoter usage, Cross-references Manual annotation alternative initiation… to over 125 databases Keywords and Gene Ontology

UniProtKB/Swiss-Prot www.uniprot.org

UniProtKB/Swiss-Prot

Manual annotation

1. Protein sequence (merge available CDS, annotate sequence discrepancies, report mistakes…)

2. Biological information (sequence analysis, extract literature information, ortholog data propagation, …)

UniProtKB/Swiss-Prot

1- Protein sequence curation

UniProtKB/Swiss-Prot

a gene-centric view of the protein space

1 entry <-> 1 gene (1 species)

The displayed protein sequence: …canonical, representative, consensus… + alternative sequences (described within the entry) What is the current status?

• At least 20% of Swiss-Prot entries required a minimal amount of curation effort so as to obtain the “correct” sequence.

• Typical problems – unsolved conflicts – uncorrected initiation sites – frameshifts – wrong gene prediction – other „problems‟

UCSC genome browser examples of CDS annotation submitted to INSDC… UniProtKB/Swiss-Prot

2- Biological data curation Extract literature information and protein sequence analysis maximum usage of controlled vocabulary

UniProtKB/Swiss-Prot gathers data form multiple sources:

- publications (literature/Pubmed) - prediction programs (Prosite, TMHMM, …) - contacts with experts - other databases - nomenclature committees

An evidence attribution system allows to easily trace the source of each annotation

Protein and gene names

General annotation (Comments)

…enable researchers to obtain a summary of what is known about a protein…

www.uniprot.org Human protein manual annotation: some statistics (June 2011)

Sequence annotation (Features)

…enable researchers to obtain a summary of what is known about a protein…

www.uniprot.org Non-experimental qualifiers

UniProtKB/Swiss-Prot considers both experimental and predicted data and makes a clear distinction between both

Type of evidence Qualifier Strong experimental evidence None or Ref.X

Light experimental evidence Probable

Inferred by similarity with homologous protein By similarity

Inferred by prediction Potential

Find all the proteins localized in the cytoplasm (experimentally proven) which are phosphorylated on a (experimentally proven) ‘Protein existence’ tag

• The „Protein existence‟ tag indicates what is the evidence for the existence of a given protein;

• Different qualifiers: 1. Evidence at protein level (~18%) (MS, western blot (tissue specificity), immuno (subcellular location),…) 2. Evidence at transcript level (~19%) 3. Inferred from homology (~58 %) 4. Predicted (~5%) 5. Uncertain (mainly in TrEMBL)

http://www.uniprot.org/docs/pe_criteria

UniProtKB

Additional information can be found in the cross-references (to more than 140 databases) Organism-specific Sequence Proteomic Genome annotation Polymorphism Family and domain AGD EMBL PeptideAtlas Ensembl dbSNP Gene3D ArachnoServer IPI PRIDE EnsemblBacteria HAMAP CGD PIR ProMEX EnsemblFungi InterPro ConoServer RefSeq EnsemblMetazoa PANTHER CTD UniGene EnsemblPlants CYGD EnsemblProtists PIRSF dictyBase GeneID PRINTS EchoBASE Gene expression GenomeReviews ProDom EcoGene KEGG PROSITE ArrayExpress euHCVdb NMPDR SMART Bgee EuPathDB TIGR SUPFAM CleanEx FlyBase UCSC TIGRFAMs Genevestigator GeneCards VectorBase GermOnline GeneDB_Spombe Protein family/group GeneFarm GenoList Allergome CAZy Gramene Ontologies H-InvDB MEROPS HGNC GO PeroxiBase HPA PptaseDB LegioList REBASE Leproma UniProtKB/Swiss-Prot: TCDB MaizeGDB MGI 129 explicit links 2D gel MIM neXtProt 2DBase-Ecoli Orphanet and 14 implicit links! ANU-2DPAGE PharmGKB Aarhus/Ghent-2DPAGE (no server) PseudoCAP COMPLUYEAST-2DPAGE RGD Cornea-2DPAGE SGD DOSAC-COBS-2DPAGE TAIR ECO2DBASE (no server) TubercuList OGP WormBase PHCI-2DPAGE Xenbase PMMA-2DPAGE ZFIN Rat-heart-2DPAGE REPRODUCTION-2DPAGE Phylogenomic dbs Siena-2DPAGE SWISS-2DPAGE eggNOG UCD-2DPAGE GeneTree World-2DPAGE HOGENOM 3D structure HOVERGEN DisProt Other PPI Enzyme and pathway InParanoid PTM HSSP OMA BindingDB DIP PDB BioCyc OrthoDB GlycoSuiteDB DrugBank IntAct PDBsum BRENDA PhylomeDB PhosphoSite NextBio MINT ProteinModelPortal Pathway_Interaction_DB ProtClustDB PhosSite PMAP-CutDB STRING SMR Reactome The UniProt web site www.uniprot.org

• Powerful search engine, google-like and easy-to-use, but also supports very directed field searches

• Scoring mechanism presenting relevant matches first

• Entry views, search result views and downloads are customizable

• The URL of a result page reflects the query; all pages and queries are bookmarkable, supporting programmatic access

• Search, Blast, Align, Retrieve, ID mapping

Search

A very powerful text search tool with autocompletion and refinement options allowing to look for UniProt entries and documentation by biological information Find all human proteins located in the nucleus The search interface guides users with helpful suggestions and hints

Advanced Search

A very powerful search tool

To be used when you know in which entry section the information is stored

Find all the protein localized in the cytoplasm (experimentally proven) which are phosphorylated on a serine (experimentally proven) Result pages: highly customizable

Result pages: downloadable

The URL can be bookmarked and manually modified. Blast

A tool associated with the standard options to search sequences in different UniProt databases and data sets Blast: customize the result display

Blast: local alignment sequence annotation highlighting option

Align

A ClustalW multiple alignment tool with sequence annotation highlighting option Align sequence annotation highlighting option

Retrieve

A UniProt specific tool allowing to retrieve a list of entries in several standard identifiers formats. You can then query your „personal database‟ with the UniProt search tool.

Query your own dataset ID Mapping

Gives the possibility to get a mapping between different databases for a given protein

These identifiers are all pointing to a TP53 (p53) protein sequence !

P04637, NP_000537, NP_001119584.1, NP_001119585.1, NP_001119584.1, NP_001119584.1, NP_001119584.1, NP_001119584.1, ENSG00000141510, CCDS11118, UPI000002ED67, IPI00025087, etc.

Download Download UniProt http://www.uniprot.org/downloads

Canonical and isoform sequences ( format) A few words on the UniProt ‘complete proteome’ sequence sets…

UniProtKB - complete proteomes

2’747 complete proteomes

 Genome completely sequenced

 Proteins mapped to the genome

 Entries tagged with the KW „Complete proteome‟

 UniProtKB/Swiss-Prot isoform sequences are available in FASTA format only

Fully manually reviewed (e.g. S. cerevisiae) Partially manually reviewed (e.g. Homo sapiens) Unreviewed (e.g. Acinetobacter baumannii (strain 1656-2)) UniProtKB - complete proteomes

Can be downloaded:

 From our complete proteome page www.uniprot.org/taxonomy/complete-proteomes

 From the „ftp download „ page

 By querying UniProtKB + download Query: organism:93062 AND keyword:"complete proteome"

Additional information: www.uniprot.org/faq/15 Query UniProtKB + download

Human proteome ~ 20‟200 genes

Query for „homo sapiens‟ (August 2011) • UniProtKB: 110,056 entries + alt sequences (~ 15‟435) = 125‟491 • UniProtKB/Swiss-Prot: 20‟244 entries + alt sequences (~ 15‟435) = 35‟679 • UniProtKB/TrEMBL: 89,834 entries • RefSeq: 32‟898 sequences • Ensembl: 90‟720 sequences

Query for „homo sapiens‟ + Complete proteome (KW-181) • UniProtKB: 56‟392 + alt sequences (15‟435) = 71‟827 • UniProtKB/Swiss-Prot: 20‟238 + alt sequences (15‟435) = 35‟673 • UniProtKB/TrEMBL: 36‟154

92% of human entries are linked with at least one RefSeq entry… Summary

Do not hesitate to contact us !

[email protected] The UniProt Consortium

SIB Ioannis Xenarios, Lydie Bougueleret, Andrea Auchincloss, Kristian Axelsen, Delphine Baratin, Marie- Claude Blatter, Brigitte Boeckmann, Jerven Bolleman, Laurent Bollondi, Emmanuel Boutet, Lionel Breuza, Alan Bridge, Edouard de Castro, Lorenzo Cerutti, Elisabeth Coudert, Béatrice Cuche, Mikael Doche, Dolnide Dornevil, Severine Duvaud, Anne Estreicher, Livia Famiglietti, Marc Feuermann, Sebastien Gehant, Elisabeth Gasteiger, Alain Gateau, Vivienne Gerritsen, Arnaud Gos, Nadine Gruaz- Gumowski, Ursula Hinz, Chantal Hulo, Nicolas Hulo, Janet James, Florence Jungo, Guillaume Keller, Vicente Lara, Philippe Lemercier, Damien Lieberherr, Xavier Martin, Patrick Masson, Anne Morgat, Salvo Paesano, Ivo Pedruzzi, Sandrine Pilbout, Sylvain Poux, Monica Pozzato, Manuela Pruess, Nicole Redaschi, Catherine Rivoire, Bernd Roechert, Michel Schneider, Christian Sigrist, Karin Sonesson, Sylvie Staehli, Eleanor Stanley, André Stutz, Shyamala Sundaram, Michael Tognolli, Laure Verbregue, Anne-Lise Veuthey

EBI Rolf Apweiler, Maria Jesus Martin, Claire O'Donovan, Michele Magrane, Yasmin Alam-Faruque, Ricardo Antunes, Benoit Bely, Mark Bingley, David Binns, Lawrence Bower, Wei Mun Chan, Emily Dimmer, Francesco Fazzini, Alexander Fedotov, John Garavelli, Leyla Garcia Castro, Rachael Huntley, Julius Jacobsen, Michael Kleen, Duncan Legge, Wudong Liu, Jie Luo, Sandra Orchard, Samuel Patient, Klemens Pichler, Diego Poggioli, Nikolas Pontikos, Steven Rosanoff, Tony Sawford, Harminder Sehra, Edward Turner, Matt Corbett, Mike Donnelly and Pieter van Rensburg

PIR Cathy H. Wu, Cecilia N. Arighi, Leslie Arminski, Winona C. Barker, Chuming Chen, Yongxing Chen, Pratibha Dubey, Hongzhan Huang, Kati Laiho, Raja Mazumder, Peter McGarvey, Darren A. Natale, Thanemozhi G. Natarajan, Jules Nchoutmboube, Natalia V. Roberts, Baris E. Suzek, Uzoamaka Ugochukwu, C. R. Vinayaka, Qinghua Wang, Yuqi Wang, Lai-Su Yeh and Jian Zhang

www.uniprot.org UniProt is mainly supported by the National Institutes of Health (NIH) grant 1 U41 HG006104- 01. Additional support for the EBI's involvement in UniProt comes from the NIH grant 2P41 HG02273-07. Swiss-Prot activities at the SIB are supported by the Swiss Federal Government through the Federal Office of Education and Science and the European Commission contracts SLING (226073), Gen2Phen (200754) and MICROME (222886). PIR activities are also supported by the NIH grants 5R01GM080646-04, 3R01GM080646-04S2, 1G08LM010720-01, and 3P20RR016472-09S2, and NSF grant DBI-0850319. www.isb-sib.ch Thank you for your attention

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