An introduction to biological databases Database or databank

An introduction to biological databases

Database or databank ? At the beginning, subtle distinctions were done between databases and databanks (in UK, but not in the USA), such as: « Database management programs for the gestion of databanks » From now on, the term « database » (db) is usually preferred

What is a database ? A collection of. . . structured searchable (index) updated periodically (release) cross-referenced (hyperlinks) -> table of contents -> new edition -> links with other db …data Includes also associated tools (software) necessary for db access, db updating, db information insertion, db information deletion….

Databases: an simple example « Introduction To Database » Teacher Database (ITDTdb) (flat file, 3 entries) Accession number: 1 First Name: Amos Last Name: Bairoch Course: DEA=oct-nov-dec 2000 http: //expasy 4. expasy. ch/people/amos. html // Accession number: 2 First Name: Laurent Last name: Falquet Course: EMBnet=sept 2000; DEA=oct-nov-dec 2000; // Accession number 3: First Name: Marie-Claude Last name: Blatter Garin Course: EMBnet=sept 2000; DEA=oct-nov-dec 2000; http: //expasy 4. expasy. ch/people/Marie-Claude. Blatter-Garin. html // Easy to manage: all the entries are visible at the same time !

Databases: an simple example (cont. ) Relational database ( « table file » ): Teacher Accession number Education Amos 1 Biochemistry Laurent 2 Biochemistry M-Claude 3 Biochemistry Course Date Involved teachers DEA Oct-nov-dec 2000 1, 3 EMBnet Sept 2000 2, 3 Easier to manage; choice of the output

Why biological databases ? Explosive growth in biological data Data (sequences, 3 D structures, 2 D gel analysis, MS analysis…. ) are no longer published in a conventional manner, but directly submitted to databases Essential tools for biological research, as classical publications used to be !

Biological databases Some databases in the field of molecular biology… AATDB, Ace. Db, ACUTS, ADB, AFDB, AGIS, AMSdb, ARR, As. Db, BBDB, BCGD, Beanref, Biolmage, Bio. Mag. Res. Bank, BIOMDB, BLOCKS, Bov. GBASE, BOVMAP, BSORF, BTKbase, CANSITE, Carb. Bank, CARBHYD, CATH, CAZY, CCDC, CD 4 OLbase, CGAP, Chick. GBASE, Colibri, COPE, Cotton. DB, CSNDB, CUTG, Cyano. Base, db. CFC, db. EST, db. STS, DDBJ, DGP, Dicty. Db, Picty_c. DB, DIP, DOGS, DOMO, DPD, DPlnteract, ECDC, ECGC, EC 02 DBASE, Eco. Cyc, Eco. Gene, EMBL, EMD db, ENZYME, EPD, Epo. DB, ESTHER, Fly. Base, Fly. View, GCRDB, GENATLAS, Genbank, Gene. Cards, Genline, Gen. Link, GENOTK, Gen. Prot. EC, GIFTS, GPCRDB, GRAP, GRBase, g. RNAsdb, GRR, GSDB, HAEMB, HAMSTERS, HEART-2 DPAGE, HEXAdb, HGMD, HIDB, HIDC, Hl. Vdb, Hot. Molec. Base, HOVERGEN, HPDB, HSC-2 DPAGE, ICN, ICTVDB, IL 2 RGbase, IMGT, Kabat, KDNA, KEGG, Klotho, LGIC, MAD, Maize. Db, MDB, Medline, Mendel, MEROPS, MGDB, MGI, MHCPEP 5 Micado, Mito. Dat, MITOMAP, MJDB, Mmt. DB, Mol-R-Us, MPDB, MRR, Mut. Base, Myc. DB, NRSub, 0 -lyc. Base, OMIA, OMIM, OPD, ORDB, OWL, PAHdb, Pat. Base, PDB, PDD, Pfam, Phospho. Base, Pig. BASE, PIR, PKR, PMD, PPDB, PRESAGE, PRINTS, Pro. Dom, Prolysis, PROSITE, PROTOMAP, Rat. MAP, RDP, REBASE, RGP, SBASE, SCOP, Seq. Anai. Ref, SGD, SGP, Sheep. Map, Soybase, SPAD, SRNA db, SRPDB, STACK, Sty. Gene, Sub 2 D, Subti. List, SWISS-2 DPAGE, SWISS-3 DIMAGE, SWISSMODEL Repository, SWISS-PROT, Tel. DB, TGN, tm. RDB, TOPS, TRANSFAC, TRR, Uni. Gene, URNADB, V BASE, VDRR, Vector. DB, WDCM, WIT, Worm. Pep, YEPD, YPM, etc. . . . !!!!

Some statistics More than 1000 different databases Generally accessible through the web (useful link: www. expasy. ch/alinks. html) Variable size: <100 Kb to >10 Gb DNA: > 10 Gb Protein: 1 Gb 3 D structure: 5 Gb Other: smaller Update frequency: daily to annually

Categories of databases for Life Sciences Sequences (DNA, protein) -> Primary db Genomics Protein domain/family -> Secondary db Mutation/polymorphism Proteomics (2 D gel, MS) 3 D structure -> Structure db Metabolism Bibliography Others

Distribution of sequence databases Books, articles Computer tapes Floppy disks CD-ROM FTP On-line services WWW DVD 1968 -> 1985 1982 ->1992 1984 -> 1990 1989 -> ? 1982 -> 1994 1993 -> ? 2001 -> ?

Sequence Databases: some « technical » definitions Data storage management: Format (flat file): fasta GCG NBRF/PIR MSF…. standardized format ? Federated databases: different autonomous, redundant, heterogeneous db linked together by links/hyperlinks. flat file: text file relational (e. g. , Oracle) object oriented (rare in biological field)

Ideal minimal content of a « sequence » db Sequences !! Accession number (AC) References Taxonomic data ANNOTATION/CURATION Keywords Cross-references Documentation

Sequence database: example SWISS-PROT Flat file taxonomy reference annotations Cross-references Keywords ID EPO_HUMAN STANDARD; PRT; 193 AA. AC P 01588; DT 21 -JUL-1986 (Rel. 01, Created) DT 21 -JUL-1986 (Rel. 01, Last sequence update) DT 30 -MAY-2000 (Rel. 39, Last annotation update) DE Erythropoietin precursor. GN EPO. OS Homo sapiens (Human). OC Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi; OC Mammalia; Eutheria; Primates; Catarrhini; Hominidae; Homo. RN [1] RP SEQUENCE FROM N. A. RX MEDLINE; 85137899. RA Jacobs K. , Shoemaker C. , Rudersdorf R. , Neill S. D. , Kaufman R. J. , RA Mufson A. , Seehra J. , Jones S. S. , Hewick R. , Fritsch E. F. , RA Kawakita M. , Shimizu T. , Miyake T. ; RT "Isolation and characterization of genomic and c. DNA clones of human RT erythropoietin. "; RL Nature 313: 806 -810(1985). . CC -!- FUNCTION: ERYTHROPOIETIN IS THE PRINCIPAL HORMONE INVOLVED IN THE CC REGULATION OF ERYTHROCYTE DIFFERENTIATION AND THE MAINTENANCE OF A CC PHYSIOLOGICAL LEVEL OF CIRCULATING ERYTHROCYTE MASS. CC -!- SUBCELLULAR LOCATION: SECRETED. CC -!- TISSUE SPECIFICITY: PRODUCED BY KIDNEY OR LIVER OF ADULT MAMMALS CC AND BY LIVER OF FETAL OR NEONATAL MAMMALS. CC -!- PHARMACEUTICAL: Available under the names Epogen (Amgen) and CC Procrit (Ortho Biotech). CC -!- DATABASE: NAME=R&D Systems' cytokine source book; CC WWW="http: //www. rndsystems. com/cyt_cat/epo. html". DR EMBL; X 02158; CAA 26095. 1; -. DR EMBL; X 02157; CAA 26094. 1; -. DR EMBL; M 11319; AAA 52400. 1; -. DR EMBL; AF 053356; AAC 78791. 1; -. DR EMBL; AF 202308; AAF 23132. 1; -. DR EMBL; AF 202306; AAF 23132. 1; JOINED. . KW Erythrocyte maturation; Glycoprotein; Hormone; Signal; Pharmaceutical. FT SIGNAL 1 27 FT CHAIN 28 193 ERYTHROPOIETIN. FT PROPEP 190 193 MAY BE REMOVED IN PROCESSED PROTEIN. FT DISULFID 34 188. . .

Sequence database: example (cont. ) sequence FT DISULFID 34 188 FT DISULFID 56 60 FT CARBOHYD 51 51 N-LINKED (GLCNAC. . . ). FT CARBOHYD 65 65 N-LINKED (GLCNAC. . . ). FT CARBOHYD 110 N-LINKED (GLCNAC. . . ). FT CARBOHYD 153 FT CONFLICT 40 40 E -> Q (IN CAA 26095). FT CONFLICT 85 85 Q -> QQ (IN REF. 5). FT CONFLICT 140 G -> R (IN CAA 26095). ** Chromosomal location: 7 q 22 SQ SEQUENCE 193 AA; 21306 MW; C 91 F 0 E 4 C 26 A 52033 CRC 64; MGVHECPAWL WLLLSLLSLP LGLPVLGAPP RLICDSRVLE RYLLEAKEAE NITTGCAEHC SLNENITVPD TKVNFYAWKR MEVGQQAVEV WQGLALLSEA VLRGQALLVN SSQPWEPLQL HVDKAVSGLR SLTTLLRALG AQKEAISPPD AASAAPLRTI TADTFRKLFR VYSNFLRGKL KLYTGEACRT GDR //

Sequence database: example …a SWISS-PROT entry, in fasta format: >sp|P 01588|EPO_HUMAN ERYTHROPOIETIN PRECURSOR - Homo sapiens (Human). MGVHECPAWLWLLLSLLSLPLGLPVLGAPPRLICDSRVLERYLLEAKEAE NITTGCAEHCSLNENITVPDTKVNFYAWKRMEVGQQAVEVWQGLALLSEA VLRGQALLVNSSQPWEPLQLHVDKAVSGLRSLTTLLRALGAQKEAISPPD AASAAPLRTITADTFRKLFRVYSNFLRGKLKLYTGEACRTGDR

Databases 1: nucleotide sequence The main DNA sequence db are EMBL (Europe)/Gen. Bank (USA) /DDBJ (Japan) There also specialized databases for the different types of RNAs (i. e. t. RNA, r. RNA, tm RNA, u. RNA, etc…) 3 D structure (DNA and RNA) Others: Aberrant splicing db; Eucaryotic promoter db (EPD); RNA editing sites, Multimedia Telomere Resource ……

EMBL/Gen. Bank/DDJB These 3 db contain mainly the same informations within 2 -3 days (few differences in the format and syntax) Serve as archives containing all sequences (single genes, ESTs, complete genomes, etc. ) derived from: Genome projects and sequencing centers Individual scientists Patent offices (i. e. European Patent Office, EPO) Non-confidential data are exchanged daily Currently: 8. 3 x 106 sequences, over 9. 7 x 109 bp; Sequences from > 50’ 000 different species;

EMBL/Gen. Bank/DDBJ Heterogeneous sequence length: genomes, variants, fragments… Sequence sizes: max 300’ 000 bp /entry (! genomic sequences, overlapping) min 10 bp /entry Archive: nothing goes out -> highly redundant ! full of errors: in sequences, in annotations, in CDS attribution… no consistency of annotations; most annotations are done by the submitters; heterogeneity of the quality and the completion and updating of the informations

EMBL/Gen. Bank/DDJB Unexpected informations you can find in these db: FT source 1. . 124 FT /db_xref="taxon: 4097" FT /organelle="plastid: chloroplast" FT /organism="Nicotiana tabacum" FT /isolate="Cuban cahibo cigar, gift from President Fidel FT Castro" Or: FT source 1. . 17084 FT /chromosome="complete mitochondrial genome" FT /db_xref="taxon: 9267" FT /organelle="mitochondrion" FT /organism="Didelphis virginiana" FT /dev_stage="adult" FT /isolate="fresh road killed individual" FT /tissue_type="liver"

EMBL entry: example ID HSERPG standard; DNA; HUM; 3398 BP. XX AC X 02158; XX SV X 02158. 1 XX DT 13 -JUN-1985 (Rel. 06, Created) DT 22 -JUN-1993 (Rel. 36, Last updated, Version 2) XX DE Human gene for erythropoietin XX KW erythropoietin; glycoprotein hormone; signal peptide. XX OS Homo sapiens (human) OC Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi; Mammalia; OC Eutheria; Primates; Catarrhini; Hominidae; Homo. XX RN [1] RP 1 -3398 RX MEDLINE; 85137899. RA Jacobs K. , Shoemaker C. , Rudersdorf R. , Neill S. D. , Kaufman R. J. , RA Mufson A. , Seehra J. , Jones S. S. , Hewick R. , Fritsch E. F. , Kawakita M. , RA Shimizu T. , Miyake T. ; RT Isolation and characterization of genomic and c. DNA clones of human RT erythropoietin; RL Nature 313: 806 -810(1985). XX DR GDB; 119110; EPO. DR GDB; 119615; TIMP 1. DR SWISS-PROT; P 01588; EPO_HUMAN. XX … keyword taxonomy references Cross-references

EMBL entry (cont. ) CC Data kindly reviewed (24 -FEB-1986) by K. Jacobs FH Key Location/Qualifiers FH FT source 1. . 3398 FT /db_xref=taxon: 9606 FT /organism=Homo sapiens FT m. RNA join(397. . 627, 1194. . 1339, 1596. . 1682, 2294. . 2473, 2608. . 3327) FT CDS join(615. . 627, 1194. . 1339, 1596. . 1682, 2294. . 2473, 2608. . 2763) FT /db_xref=SWISS-PROT: P 01588 FT /product=erythropoietin FT /protein_id=CAA 26095. 1 FT /translation=MGVHECPAWLWLLLSLLSLPLGLPVLGAPPRLICDSRVLQRYLLE FT AKEAENITTGCAEHCSLNENITVPDTKVNFYAWKRMEVGQQAVEVWQGLALLSEAVLRG FT QALLVNSSQPWEPLQLHVDKAVSGLRSLTTLLRALGAQKEAISPPDAASAAPLRTITAD FT TFRKLFRVYSNFLRGKLKLYTGEACRTGDR FT mat_peptide join(1262. . 1339, 1596. . 1682, 2294. . 2473, 2608. . 2763) FT /product=erythropoietin FT sig_peptide join(615. . 627, 1194. . 1261) FT exon 397. . 627 FT /number=1 FT intron 628. . 1193 FT /number=1 FT exon 1194. . 1339 FT /number=2 FT intron 1340. . 1595 FT /number=2 FT exon 1596. . 1682 FT /number=3 FT intron 1683. . 2293 FT /number=3 FT exon 2294. . 2473 FT /number=4 FT intron 2474. . 2607 FT /number=4 FT exon 2608. . 3327 FT /note=3' untranslated region FT /number=5 XX SQ Sequence 3398 BP; 698 A; 1034 C; 991 G; 675 T; 0 other; agcttctggg cttccagacc cagctacttt gcggaactca gcaacccagg catctctgag 60 tctccgccca agaccgggat gccccccagg aggtgtccgg gagcccagcc tttcccagat 120 annotation sequence

Gen. Bank entry: example LOCUS HSERPG 3398 bp DNA PRI 22 -JUN-1993 DEFINITION Human gene for erythropoietin. ACCESSION X 02158 VERSION X 02158. 1 GI: 31224 KEYWORDS erythropoietin; glycoprotein hormone; signal peptide. SOURCE human. ORGANISM Homo sapiens Eukaryota; Metazoa; Chordata; Vertebrata; Mammalia; Eutheria; Primates; Catarrhini; Hominidae; Homo. REFERENCE 1 (bases 1 to 3398) AUTHORS Jacobs, K. , Shoemaker, C. , Rudersdorf, R. , Neill, S. D. , Kaufman, R. J. , Mufson, A. , Seehra, J. , Jones, S. S. , Hewick, R. , Fritsch, E. F. , Kawakita, M. , Shimizu, T. and Miyake, T. TITLE Isolation and characterization of genomic and c. DNA clones of human erythropoietin JOURNAL Nature 313 (6005), 806 -810 (1985) MEDLINE 85137899 COMMENT Data kindly reviewed (24 -FEB-1986) by K. Jacobs. FEATURES Location/Qualifiers source 1. . 3398 /organism="Homo sapiens" /db_ xref="taxon: 9606" m. RNA join(397. . 627, 1194. . 1339, 1596. . 1682, 2294. . 2473, 2608. . 3327) exon 397. . 627 /number=1 sig_peptide join(615. . 627, 1194. . 1261) CDS join(615. . 627, 1194. . 1339, 1596. . 1682, 2294. . 2473, 2608. . 2763) / codon_start=1 /product=" erythropoietin" /protein_id="CAA 26095. 1" / db_xref="GI: 312304" / db_xref="SWISS-PROT: P 01588" /translation="MGVHECPAWLWLLLSLLSLPLGLPVLGAPPRLICDSRVLQRYLL EAKEAENITTGCAEHCSLNENITVPDTKVNFYAWKRMEVGQQAVEVWQGLALLSEAVL RGQALLVNSSQPWEPLQLHVDKAVSGLRSLTTLLRALGAQKEAISPPDAASAAPLRTI …

Gen. Bank entry (cont. ) TADTFRKLFRVYSNFLRGKLKLYTGEACRTGDR" intron 628. . 1193 /number=1 exon 1194. . 1339 /number=2 mat_peptide join(1262. . 1339, 1596. . 1682, 2294. . 2473, 2608. . 2760) /product="erythropoietin" intron 1340. . 1595 /number=2 exon 1596. . 1682 /number=3 intron 1683. . 2293 /number=3 exon 2294. . 2473 /number=4 intron 2474. . 2607 /number=4 exon 2608. . 3327 /note="3' untranslated region" /number=5 BASE COUNT 698 a 1034 c 991 g 675 t ORIGIN 1 agcttctggg cttccagacc cagctacttt gcggaactca gcaacccagg catctctgag 61 tctccgccca agaccgggat gccccccagg aggtgtccgg gagcccagcc tttcccagat 121 agcagctccg ccagtcccaa gggtgcgcaa ccggctgcac tcccctcccg cgacccaggg 181 cccgggagca gcccccatga cccacacgca cgtctgcagc agccccgtca gccccggagc 241 ctcaacccag gcgtcctgcc cctgctctga ccccgggtgg cccctacccc tggcgacccc

DDJB entry: example LOCUS HSERPG 3398 bp DNA HUM 22 -JUN-1993 DEFINITION Human gene for erythropoietin. ACCESSION X 02158 VERSION X 02158. 1 KEYWORDS erythropoietin; glycoprotein hormone; signal peptide. SOURCE human. ORGANISM Homo sapiens Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Mammalia; Eutheria; Primates; Catarrhini; Hominidae; Homo. REFERENCE 1 (bases 1 to 3398) AUTHORS Jacobs, K. , Shoemaker, C. , Rudersdorf, R. , Neill, S. D. , Kaufman, R. J. , Mufson, A. , Seehra, J. , Jones, S. S. , Hewick, R. , Fritsch, E. F. , Kawakita, M. , Shimizu, T. and Miyake, T. TITLE Isolation and characterization of genomic and c. DNA clones of human erythropoietin JOURNAL Nature 313, 806 -810(1985) MEDLINE 85137899 COMMENT Data kindly reviewed (24 -FEB-1986) by K. Jacobs FEATURES Location/Qualifiers source 1. . 3398 /db_xref="taxon: 9606" /organism="Homo sapiens" m. RNA join(397. . 627, 1194. . 1339, 1596. . 1682, 2294. . 2473, 2608. . 3327) CDS join(615. . 627, 1194. . 1339, 1596. . 1682, 2294. . 2473, 2608. . 2763) /db_xref="SWISS-PROT: P 01588" /product="erythropoietin" /protein_id="CAA 26095. 1" /translation="MGVHECPAWLWLLLSLLSLPLGLPVLGAPPRLICDSRVLQRYLLE AKEAENITTGCAEHCSLNENITVPDTKVNFYAWKRMEVGQQAVEVWQGLALLSEAVLRG QALLVNSSQPWEPLQLHVDKAVSGLRSLTTLLRALGAQKEAISPPDAASAAPLRTITAD TFRKLFRVYSNFLRGKLKLYTGEACRTGDR » …

DDJB (cont. ) mat_peptide join(1262. . 1339, 1596. . 1682, 2294. . 2473, 2608. . 2763) /product="erythropoietin" sig_peptide join(615. . 627, 1194. . 1261) exon 397. . 627 /number=1 intron 628. . 1193 /number=1 exon 1194. . 1339 /number=2 intron 1340. . 1595 /number=2 exon 1596. . 1682 /number=3 intron 1683. . 2293 /number=3 exon 2294. . 2473 /number=4 intron 2474. . 2607 /number=4 exon 2608. . 3327 /note="3' untranslated region" /number=5 BASE COUNT 698 a 1034 c 991 g 675 t ORIGIN 1 agcttctggg cttccagacc cagctacttt gcggaactca gcaacccagg catctctgag 61 tctccgccca agaccgggat gccccccagg aggtgtccgg gagcccagcc tttcccagat

The tremendous increase in nucleotide sequences EMBL data…first increase in data due to the PCR development… 1980: 80 genes fully sequenced !

EMBL divisions EMBL has been divided into subdatabases to allow easier data management and searches fun, hum, inv, mam, org, phg, pln, pro, rod, syn, unc, vrl, vrt est, gss, htg, sts, patent

Ref. Seq a SWISS-PROT clone? The NCBI Reference Sequence project (Ref. Seq) will provide reference sequence standards for the naturally occurring molecules of the central dogma, from chromosomes to m. RNAs to proteins. Ref. Seq standards provide a foundation for the functional annotation of the human genome. They provide a stable reference point for mutation analysis, gene expression studies, and polymorphism discovery. Molecule Accession Format Genome Complete Genome NC_###### Archaea, Bacterial, Organelle, Virus, Viroid Complete Chrom. NC_###### Eukaryote Complete Sequence NC_###### Plasmid Genomic Contig NT_###### Homo sapiens m. RNA NM_###### Homo sapiens, Mus musculus, Rattus norvegicus Protein NP_###### All of the above

Ref. Seq a SWISS-PROT clone? Ref. Seq records are created via a process consisting of: identifying sequences that represent distinct genes establishing the correct gene name-to-accession number association identifying the full extent of available sequence data creating a new Ref. Seq record with a status of: PREDICTED PROVISIONAL REVIEWED Provisional Ref. Seq records are reviewed by a biologist who confirms the initial name-to-sequence association, adds information including a summary of gene function, and, more importantly, corrects, re-annotates, or extends the sequence data using data available in other Gen. Bank records.

Databases 2: genomics Contain information on genes, gene location (mapping), gene nomenclature and links to sequence databases; Exist for most organisms important for life science research; Examples: MIM, GDB (human), MGD (mouse), Fly. Base (Drosophila), SGD (yeast), Maize. DB (maize), Subti. List (B. subtilis), etc. ; Format: generally relational (Oracle, Sy. Base or Ace. Db).

MIM OMIM™: Online Mendelian Inheritance in Man a catalog of human genes and genetic disorders contains a summary of literature, pictures, and reference information. It also contains numerous links to articles and sequence information.

MIM: example *133170 ERYTHROPOIETIN; EPO Alternative titles; symbols EP TABLE OF CONTENTS TEXT REFERENCES SEE ALSO CONTRIBUTORS CREATION DATE EDIT HISTORY Database Links Gene Map Locus: 7 q 21 Note: pressing the symbol will find the citations in MEDLINE whose text most closely matches the text of the preceding OMIM paragraph, using the Entrez MEDLINE neighboring function. TEXT Human erythropoietin is an acidic glycoprotein hormone with molecular weight 34, 000. As the prime regulator of red cell production, its major functions are to promote erythroid differentiation and to initiate hemoglobin synthesis. Sherwood and Shouval (1986) described a human renal carcinoma cell line that continuously produces erythropoietin. Eschbach et al. (1987) demonstrated the effectiveness of recombinant human erythropoietin in treating the anemia of end-stage renal disease. Lee-Huang (1984) cloned human erythropoietin c. DNA in E. coli. Mc. Donald et al. (1986) and Shoemaker and Mitsock (1986) cloned the mouse gene and the latter workers showed that coding DNA and amino acid sequence are about 80% conserved between man and mouse. This is a much higher order of conservation than for various interferons, interleukin-2, and GM-CSF. ……

Ensembl Contains all the human genome DNA sequences currently available in the public domain. Automated annotation: by using different software tools, features are identified in the DNA sequences: Genes (known or predicted) Single nucleotide polymorphisms (SNPs) Repeats Homologies Created and maintained by the EBI and the Sanger Center (UK) www. ensembl. org

Database 3: protein sequence SWISS-PROT: created in 1986 (A. Bairoch) Tr. EMBL: created in 1996; complement to SWISS-PROT; derived Gen. Pept: derived from automated Gen. Bank CDS translations and from automated EMBL CDS translations ( « proteomic » version of EMBL) journal scans ( « proteomic » version of Gen. Bank) PIR: Protein Information Resources MIPS: Martinsried Institute for Protein Sequences PIR + PATCHX (supplement of unverified protein sequences from external sources)

Database 3: protein sequence NRL-3 D: produced by PIR from PDB (3 D struture) sequences Many specialized protein databases for specific families or groups of proteins. Examples: YPD (yeast proteins), AMSDb (antibacterial peptides), GPCRDB (7 TM receptors), IMGT (immune system) etc.

SWISS-PROT Collaboration between the SIB (CH) and EMBL/EBI (UK) Annotated (manually), non-redundant, crossreferenced, documented protein sequence database. 88 ’ 000 sequences from more than 6’ 800 different species; 70 ’ 000 references (publications); 550 ’ 000 cross-references (databases); ~200 Mb of annotations. Weekly releases; available from about 50 servers across the world, the main source being Ex. PASy

SWISS-PROT: example Never changed

SWISS-PROT (cont. )

Tr. EMBL (Translation of EMBL) Computer-annotated supplement to SWISS-PROT, as it is impossible to cope with the flow of data… Well-structure SWISS-PROT-like resource Derived from automated EMBL CDS translation (maintained at the EBI (UK)) Tr. EMBL is automatically generated annotated using software tools (incompatible with the SWISS-PROT in terms of quality) Tr. EMBL contains all what is not yet in SWISSPROT Yerk!! But there is no choice and these software tools are becoming quite good !

The simplified story of a Sprot entry c. DNAs, genomes, …. EMBLnew EMBL « Automatic » • Redundancy check (merge) • Inter. Pro (family attribution) • Annotation Tr. EMBL « Manual » • Redundancy (merge, conflicts) • Annotation • Sprot tools (macros…) • Sprot documentation • Medline • Databases (MIM, MGD…. ) • Brain storming CDS Tr. EMBLnew SWISS-PROT Once in Sprot, the entry is no more in Tr. EMBL, but still in EMBL (archive)

SWISS-PROT introduces a new arithmetical concept ! How many sequences in SWISS-PROT + Tr. EMBL ? 88’ 000 + 300 ’ 000 = about 240’ 000 SWISS-PROT and Tr. EMBL (SPTR) a minimal of redundancy

Tr. EMBL divisions Tr. EMBL: SPTr. EMBL + REMTr. EMBL SPTr. EMBL: Tr. EMBL entries that will eventually be integrated into SWISS-PROT, but that have not yet be manually annotated REMTr. EMBL: sequences that are not destined to be included in SWISS-PROT Immunoglobulins and T-cell receptors Synthetic sequences Patented sequences Small fragments (<8 aa) CDS not coding for real proteins Tr. EMBL new: updates to the latest release of TREMBL

Tr. EMBL divisions Subdivisions Archae Fungus Human Invertebrate Mammals Major Hist. Comp. Organelles Phage Plant Prokaryote Rodent Uncommented Viral Vertebrate arc fun hum inv mam mhc org phg pln pro rod unc vrl vrt

Tr. EMBL: example

Gen. Pept (translation of Gen. Bank) Gen. Pept is a protein database translated from the last release of Gen. Bank (+ journal scans) The current release has 484’ 496 entries In contrast to Tr. EMBL, keeps all protein sequences including small fragments (< 8 aa), immunoglobulins…. Redundancy: 20 entries for human EPO

Gen. Pept: example LOCUS L 33410_1 [HUMMLCMPL] DEFINITION Human c- mpl ligand (ML) m. RNA, complete cds; erythropoietin homology domain bp 66. . 522. DATE 07 -JAN-1995 ACCESSION L 33410 NID ORGANISM Homo_SP_sapiens Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi; Mammalia; Eutheria; Primates; Catarrhini; Hominidae; Homo. COMMENT CDS 216. . 1277 /gene="ML" /product="c- mpl ligand" /protein_id="AAA 59857. 1" / db_xref="GI: 506827" WEIGHT 37823 LENGTH 353 ORIGIN 1 MELTELLLVV MLLLTARLTL SSPAPPACDL RVLSKLLRDS HVLHSRLSQC PEVHPLPTPV 61 LLPAVDFSLG EWKTQMEETK AQDILGAVTL LLEGVMAARG QLGPTCLSSL LGQLSGQVRL 121 LLGALQSLLG TQLPPQGRTT AHKDPNAIFL SFQHLLRGKV RFLMLVGGST LCVRRAPPTT 181 AVPSRTSLVL TLNELPNRTS GLLETNFTAS ARTTGSGLLK WQQGFRAKIP GLLNQTSRSL 241 DQIPGYLNRI HELLNGTRGL FPGPSRRTLG APDISSGTSD TGSLPPNLQP GYSPSPTHPP 301 TGQYTLFPLP PTLPTPVVQL HPLLPDPSAP TPTPTSPLLN TSYTHSQNLS QEG //

PIR Protein Information Resource, created in 1984 Successor of the National Biochemical Research Foundation (NBRF) protein sequence database developed in 1965 by M. O. Dayhoff « Atlas of Protein Sequence and Structure » Maintained by MIPS (Germany) and JIPID (Japan) Provides some cross-referencing to EMBL/Gen. Bank/DDJB and PDB, GDB, Fly. Base, OMIM, SGD, and MGD In august 2000: 178’ 050 entries. Redundancy: 3 entries for human EPO

PIR: example >P 1; ZUHU erythropoietin precursor - human C; Species: Homo sapiens (man) C; Date: 27 -Nov-1985 #sequence_revision 27 -Nov-1985 #text_change 22 -Jun-1999 C; Accession: A 01855; A 24744; A 25384; A 22210; S 56178 R; Jacobs, K. ; Shoemaker, C. ; Rudersdorf, R. ; Neill, S. D. ; Kaufman, R. J. ; Mufson, A. ; Seehra, J. ; Jones, S. S. ; Hewick, R. ; Fritsch, E. F. ; Kawakita, M. ; Shimizu, T. ; Miyake, T. Nature 313, 806 -810, 1985 A; Title: Isolation and characterization of genomic and c. DNA clones of human erythropoietin. A; Reference number: A 01855; MUID: 85137899 A; Accession: A 01855 A; Molecule type: m. RNA; DNA A; Residues: 1 -193 A; Cross-references: GB: X 02157; GB: X 02158 R; Lin, F. K. ; Suggs, S. ; Lin, C. H. ; Browne, J. K. ; Smalling, R. ; Egrie, J. C. ; Chen, K. K. ; Fox, G. M. ; Martin, F. ; Stabinsky, Z. ; Badrawi, S. M. ; Lai, P. H. ; Goldwasser, E. Proc. Natl. Acad. Sci. U. S. A. 82, 7580 -7584, 1985 A; Title: Cloning and expression of the human erythropoietin gene. A; Reference number: A 24744; MUID: 86067948 A; Accession: A 24744 A; Molecule type: DNA A; Residues: 1 -193 A; Cross-references: GB: M 11319; NID: g 182197; PIDN: AAA 52400. 1; PID: g 182198 R; Lai, P. H. ; Everett, R. ; Wang, F. F. ; Arakawa, T. ; Goldwasser, E. J. Biol. Chem. 261, 3116 -3121, 1986 A; Title: Structural characterization of human erythropoietin. A; Reference number: A 25384; MUID: 86140080 A; Accession: A 25384 A; Molecule type: protein A; Residues: 28 -86, 'Q', 87 -193 A; Experimental source: urine A; Note: forms without the carboxyl-terminal residue and the four carboxyl-terminal residues were observed R; Yanagawa, S. ; Hirade, K. ; Ohnota, H. ; Sasaki, R. ; Chiba, H. ; Ueda, M. ; Goto, M. J. Biol. Chem. 259, 2707 -2710, 1984 A; Title: Isolation of human erythropoietin with monoclonal antibodies. A; Reference number: A 22210; MUID: 84135751

PIR (cont. ) A; Accession: A 22210 A; Molecule type: protein A; Residues: 28 -29, 'X', 31 -33, 'L', 35 -50, 'X', 52 -53, 'D', 55, 'G', 57 R; Matsumoto, S. ; Ikura, K. ; Ueda, M. ; Sasaki, R. Plant Mol. Biol. 27, 1163 -1172, 1995 A; Title: Characterization of a human glycoprotein (erythropoietin) produced in cultured tobacco cells. A; Reference number: S 56178; MUID: 95284365 A; Accession: S 56178 A; Molecule type: protein A; Residues: 28 -33, 'X', 35 -37 C; Comment: Erythropoietin is produced by kidney or liver of adult mammals and by liver of fetal or neonatal mammals. C; Genetics: A; Gene: GDB: EPO A; Cross-references: GDB: 119110; OMIM: 133170 A; Map position: 7 q 21. 3 -7 q 22. 1 A; Introns: 5/1; 53/3; 82/3; 142/3 C; Function: A; Description: the primary inducer of erythrocyte formation C; Superfamily: erythropoietin C; Keywords: erythropoiesis; glycoprotein; hormone; kidney; liver F; 1 -27/Domain: signal sequence #status predicted F; 28 -193/Product: erythropoietin #status experimental F; 34 -188, 56 -60/Disulfide bonds: #status experimental F; 51, 65, 110/Binding site: carbohydrate ( Asn) (covalent) #status experimental F; 153/Binding site: carbohydrate (Ser) (covalent) #status experimental >P 1; ZUHU MGVHECPAWL WLLLSLLSLP LGLPVLGAPP RLICDSRVLE RYLLEAKEAE NITTGCAEHC SLNENITVPD TKVNFYAWKR MEVGQQAVEV WQGLALLSEA VLRGQALLVN SSQPWEPLQL HVDKAVSGLR SLTTLLRALG AQKEAISPPD AASAAPLRTI TADTFRKLFR VYSNFLRGKL KLYTGEACRT GDR*

Composite protein sequence db Different composite db use different primary sources and different redundancy criteria in their amalgamation procedures NRDB OWL MIPSX SPTr. EMBL * PDB SWISS-PROT PIR Gen. Pept SP update Gen. Pept update SWISS-PROT PIR Gen. Bank NRL-3 D PIR MIPS NRL-3 D SWISS-PROT EMBL translation Gen. Bank translation Kabat (immuno) Pseq. IP SWISS-PROT SPTr. EMBLnew Redundancy priority criteria * Also called SWall at EBI SWIR: SPTr. EMBL + Wormpep

Composite: protein family The proteins /genes are classified by superfamily/family according to Blast/Fasta (homology) results General: Prot. Fam: PIR Proto. Map: SWISS-PROT SYSTERS: SWISS-PROT and PIR (non redundant) Pro. Class: PIR and PROSITE Species specific: HOVERGEN: vertebrates HOBACGEN: bacteria COG: complete organism genome

Proto. Map: example

Proto. Map (cont. )

Database 4: protein domain/family Contains biologically significant « pattern / profiles/ HMM » formulated in such a way that, with appropriate computional tools, it can rapidly and reliably determine to which known family of proteins (if any) a new sequence belongs to -> tools to identify what is the function of uncharacterized proteins translated from genomic or c. DNA sequences ( « functional diagnostic » )

Protein domain/family Most proteins have « modular » structure Estimation: ~ 3 domains / protein Domains (conserved sequences or structures) are identified by multi sequence alignments Domains can be defined by different methods: Pattern (regular expression); used for very conserved domains Profiles (weighted matrices): two-dimensional tables of position specific match-, gap-, and insertion-scores, derived from aligned sequence families; used for less conserved domains Hidden Markov Model (HMM); probabilistic models; an other method to generate profiles.

Some statistics 15 most common protein domains for H. sapiens (Incomplete) Immunoglobulin and major histocompatibility complex domain Eukaryotic protein kinase Zinc finger, C 2 H 2 type Rhodopsin-like GPCR superfamily Src homology 3 (SH 3) domain RNA-binding region RNP-1 (RNA recognition motif) Fibronectin type III domain Pleckstrin homology (PH) domain Homeobox domain Major histocompatibility complex protein, Class I EF-hand family EGF-like domain RING finger Cadherin domain PDZ domain (also known as DHR or GLGF) Serine proteases, trypsin family http: //www. ebi. ac. uk/proteome/HUMAN/interpro/top 15 d. html

Protein domain/family db Secondary databases are the fruit of analyses of the sequences found in the primary db Either manually curated (i. e. PROSITE, Pfam, etc. ) or automatically generated (i. e. Pro. Dom, DOMO) Some depend on the method used to detect if a protein belongs to a particular domain/family (patterns, profiles, HMM)

Protein domain/family db Secondary db Primary source Information PROSITE SWISS-PROT Patterns (Regular expression) PROSITE SWISS-PROT Profiles (Weighted matrices) PRINTS OWL and SWISS-PROT Aligned motifs Pfam SWISS-PROT HMM (Fingerprints) (Hidden Markov Models) BLOCKS PROSITE/PRINTS Aligned motifs IDENTIFY BLOCKS/PRINTS Fuzzy regular expressions

Prosite § Created in 1988 (SIB) § Contains functional domains fully annotated, based on two methods: patterns and profiles § Entries are deposited in PROSITE in two distinct files: § Pattern/profiles with the lists of all matches in the parent version of SWISS-PROT § Documentation Aug 2000: contains 1064 documentation entries that describe 1424 different patterns, rules and profiles/matrices.

Prosite (pattern): example ID AC DT DE PA NR Diagnostic NR performance NR CC CC DR List of DR matches DR DR DO // EPO_TPO; PATTERN. PS 00817; OCT-1993 (CREATED); NOV-1995 (DATA UPDATE); JUL-1998 (INFO UPDATE). Erythropoietin / thrombopoeitin signature. P-x(4)-C-D-x-R-[LIVM](2)-x-[KR]-x(14)-C. /RELEASE=38, 80000; /TOTAL=14(14); /POSITIVE=14(14); /UNKNOWN=0(0); /FALSE_POS=0(0); /FALSE_NEG=0; /PARTIAL=1; /TAXO-RANGE=? ? E? ? ; /MAX-REPEAT=1; /SITE=3, disulfide; /SITE=11, disulfide; P 48617, EPO_BOVIN , T; P 33707, EPO_CANFA , T; P 33708, EPO_FELCA , T; P 01588, EPO_HUMAN , T; P 07865, EPO_MACFA , T; Q 28513, EPO_MACMU , T; P 07321, EPO_MOUSE , T; P 49157, EPO_PIG , T; P 29676, EPO_RAT , T; P 33709, EPO_SHEEP , T; P 42705, TPO_CANFA , T; P 40225, TPO_HUMAN , T; P 40226, TPO_MOUSE , T; P 49745, TPO_RAT , T; P 42706, TPO_PIG , P; PDOC 00644;

Prosite (profile): example PROSITE: PS 50097 ID AC DT DE MA MA MA MA MA MA MA …. BTB; MATRIX. PS 50097; DEC-1999 (CREATED); DEC-1999 (DATA UPDATE); DEC-1999 (INFO UPDATE). BTB domain profile. /GENERAL_SPEC: ALPHABET='ABCDEFGHIKLMNPQRSTVWYZ'; LENGTH=67; /DISJOINT: DEFINITION=PROTECT; N 1=6; N 2=62; /NORMALIZATION: MODE=1; FUNCTION=LINEAR; R 1=. 9751; R 2=. 02068202; TEXT='-Log. E'; /CUT_OFF: LEVEL=0; SCORE=363; N_SCORE=8. 5; MODE=1; TEXT='!'; /CUT_OFF: LEVEL=-1; SCORE=267; N_SCORE=6. 5; MODE=1; TEXT='? '; /DEFAULT: D=-20; I=-20; B 1=-50; E 1=-50; MI=-105; MD=-105; IM=-105; DM=-105; MM=1; M 0=-2; /I: B 1=0; BI=-105; BD=-105; /M: SY='C'; M=-6, -10, 28, -14, -9, -15, -20, -14, -19, -15, -17, -14, -8, -19, -14, -15, 0, 0, -9, -32, -17, -12; /M: SY='D'; M=-16, 41, -28, 53, 15, -34, -11, -33, 0, -27, -25, 21, -11, 0, -8, 2, -6, -26, -38, -19, 7; /M: SY='V'; M=2, -23, -8, -24, -1, -24, -25, 16, -20, 7, 6, -20, -25, -23, -20, -10, -4, 24, -23, -9, -24; /M: SY='T'; M=-2, -13, -18, -19, -13, -7, -24, -19, 6, -8, -2, 1, -17, -11, -10, -1, 10, -24, -6, -13; /M: SY='L'; M=-11, -30, -22, -33, -24, 15, -32, -23, 25, -29, 35, 17, -26, -27, -23, -22, -24, -9, 16, -17, 3, -24; /M: SY='V'; M=0, -11, -18, -13, -10, -12, -20, -13, 1, -6, -4, 2, -10, -19, -6, -7, -4, -2, 8, -25, -9; /M: SY='V'; M=1, -25, -3, -29, -25, -26, 17, -22, 10, 7, -23, -25, -23, -22, -11, -3, 24, -27, -10, -25; /M: SY='D'; M=-6, 7, -26, 8, 7, -25, 6, -7, -27, 0, -23, -17, 8, -13, 0, -3, 3, -6, -23, -27, -17, 3; /I: I=-5; MI=0; IM=0; DM=-15; MD=-15; /M: SY='G'; M=-6, 8, -27, 8, -3, -27, 22, -7, -30, -8, -26, -19, 10, -14, -8, -9, 2, -9, -24, -28, -21, -6; /M: SY='K'; M=-7, -4, -23, -4, 7, -23, -13, -21, 10, -18, -9, -3, -12, 7, 9, -2, -4, -16, -25, -12, 6; /M: SY='E'; M=-8, -6, -21, -8, 1, -15, -21, -7, -10, -5, -3, -14, 0, -1, -2, -6, -26, -9, -1; /M: SY='F'; M=-12, -28, -22, -34, -26, 31, -21, 18, -26, 16, 9, -22, -27, -21, -20, -9, 14, -6, 13, -26; /M: SY='R'; M=-13, -9, -24, -10, -3, -11, -21, 7, -17, 7, -16, -4, -8, 2, 9, -9, -16, -20, -1, -2; /M: SY='A'; M=21, -15, -8, -22, -17, -10, -23, 0, -15, -5, -14, -18, -17, -19, 4, 6, 12, -24, -15, -17; /M: SY='H'; M=-15, 5, -22, 2, -1, -20, -16, 65, -26, -8, -21, -5, 15, -19, 6, -2, -11, -26, -32, 7, 0; /M: SY='K'; M=-12, -5, -29, -5, 5, -25, -18, -26, 34, -24, -9, -14, 8, 34, -8, -17, -20, -10, 5; /M: SY='A'; M=4, -12, -16, -10, -6, -18, -14, -2, -13, -1, -2, -11, -17, -12, -13, -3, 1, 2, -24, -8, -11; /M: SY='V'; M=-7, -26, -19, -31, -26, 7, -32, -24, 27, -23, 14, 11, -22, -25, -23, -22, -13, 0, 28, -19, 3, -26; /M: SY='L'; M=-10, -30, -21, 9, -30, -20, 22, -29, 47, 20, -29, -20, -29, -10, 12, -20, 0, -21; /M: SY='A'; M=18, -6, 0, -12, -8, -18, -6, -15, -10, -18, -12, -14, -8, -13, 18, 11, -5, -32, -19, -8;

Prosite (profile): example (cont. ) …… MA MA MA NR NR NR CC DR DR DR DR DO // /M: SY='T'; M=-3, 3, -16, 1, -3, -18, -12, -9, -20, -6, -19, -15, 2, -7, -6, 10, 15, -13, -27, -12, -5; /M: SY='G'; M=-1, 1, -25, 2, -9, -26, 31, -12, -32, -10, -26, -18, 4, -17, -12, -10, 1, -12, -24, -25, -22, -11; /M: SY='E'; M=-9, 3, -24, 4, 13, -25, -16, -1, -24, 13, -21, -13, 3, -9, 6, 13, -6, -20, -27, -13, 8; /M: SY='I'; M=-6, -21, -18, -25, -21, -29, -21, 14, 10, -19, -24, -17, -19, -13, -3, 19, -23, -20; /M: SY='E'; M=-4, 3, -23, 3, 4, -18, -11, -7, -1, -18, -13, 3, -9, -1, -5, 1, -4, -14, -25, -11, 1; /M: SY='I'; M=-8, -25, -23, -27, -20, 1, -30, -21, -20, 18, 12, -22, -18, -7, 16, -21, -20; /M: SY='P'; M=-6, 0, -24, 2, 1, -22, -13, -8, -21, -23, -15, 1, 14, -7, 3, 2, -19, -31, -18, -3; /M: SY='E'; M=-7, 1, -27, 4, 11, -24, -15, -4, -19, 2, -18, -11, 0, -1, 6, -1, -2, -6, -19, -25, -14, 7; /I: E 1=0; IE=-105; DE=-105; /RELEASE=39, 87397; /TOTAL=46(44); /POSITIVE=45(43); /UNKNOWN=1(1); /FALSE_POS=0(0); /FALSE_NEG=0; /PARTIAL=0; /TAXO-RANGE=? ? E? V; /MAX-REPEAT=2; O 14867, BAC 1_HUMAN, T; P 97302, BAC 1_MOUSE, T; P 97303, BAC 2_MOUSE, T; P 41182, BCL 6_HUMAN, T; P 41183, BCL 6_MOUSE, T; Q 01295, BRC 1_DROME, T; Q 01296, BRC 2_DROME, T; Q 01293, BRC 3_DROME, T; Q 28068, CALI_BOVIN, T; Q 13939, CALI_HUMAN, T; Q 08605, GAGA_DROME, T; Q 01820, GCL 1_DROME, T; P 10074, HKR 3_HUMAN, T; Q 04652, KELC_DROME, T; P 42283, LOLL_DROME, T; P 42284, LOLS_DROME, T; O 14682, PI 10_HUMAN, T; Q 05516, PLZF_HUMAN, T; O 43791, SPOP_HUMAN, T; P 42282, TTKA_DROME, T; P 17789, TTKB_DROME, T; P 21073, VA 55_VACCC, T; P 24768, VA 55_VACCV, T; P 21037, VC 02_VACCC, T; P 17371, VC 02_VACCV, T; P 32228, VC 04_SPVKA, T; P 32206, VC 13_SPVKA, T; P 21013, VF 03_VACCC, T; P 24357, VF 03_VACCV, T; P 22611, VMT 8_MYXVL, T; P 08073, VMT 9_MYXVL, T; O 43167, Y 441_HUMAN, T; Q 10225, YAZ 4_SCHPO, T; P 40560, YIA 1_YEAST, T; P 34324, YKV 2_CAEEL, T; P 34371, YLJ 8_CAEEL, T; P 34568, YNV 5_CAEEL, T; P 41886, YPT 9_CAEEL, T; Q 09563, YR 47_CAEEL, T; Q 10017, YSW 1_CAEEL, T; Q 13105, Z 151_HUMAN, T; Q 60821, Z 151_MOUSE, T; P 24278, ZN 46_HUMAN, T; Q 13829, TNP 1_HUMAN, ? ; PDOC 50097;

PRINTS Compendium of protein motif fingerprints Most protein families are characterized by several conserved motifs Fingerprint: set of motif(s) (simple or composite, such as multidomains) = signature of family membership True family members exhibit all elements of the fingerprint, while subfamily members may possess only a part

Pro. Dom consists of an automated compilation of homologous domain alignment (procedure based on PSI-BLAST searches) Updating problem ! Last Pro. Dom update: February 7, 2000 built from SWISS-PROT 38 + TREMBL +TREMBL updates - October 22, 1999

Pro. Dom: example Your query

Protein domain/family: Composite databases Example: Inter. Pro Unification of PROSITE, PRINTS, Pfam and Pro. Dom into an integrated resource of protein families, domains and functional sites; Single set of «documents» linked to the various methods; Will be used to improve the functional annotation of SWISS-PROT (classification of unknown protein…) This release contains 3052 entries, representing 574 domains, 2418 families, 46 repeats and 14 post-translational modification sites.

Inter. Pro: example IPR 001323 Name Type Erythropoietin/thrombopoeitin Family Abstract Erythropoietin, a plasma glycoprotein, is the primary physiological mediator of erythropoiesis [1]. It is involved in the regulation of the level of peripheral erythrocytes by stimulating the differentiation of erythroid progenitor cells, found in the spleen and bone marrow, into mature erythrocytes [2]. It is primarily produced in adult kidneys and foetal liver, acting by attachment to specific binding sites on erythroid progenitor cells, stimulating their differentiation [3]. Severe kidney dysfunction causes reduction in the plasma levels of erythropoietin, resulting in chronic anaemia - injection of purified erythropoietin into the blood stream can help to relieve this type of anaemia. Levels of erythropoietin in plasma fluctuate with varying oxygen tension of the blood, but androgens and prostaglandins also modulate the levels to some extent [3]. Erythropoietin glycoprotein sequences are well conserved, a consequence of which is that the hormones are cross-reactive among mammals, i. e. that from one species, say human, can stimulate erythropoiesis in other species, say mouse or rat [4]. …. Thrombopoeitin (TPO), a glycoprotein, is the mammalian hormone which functions as a megakaryocytic lineage specific growth and differentiation factor affecting the proliferation and maturation from their committed progenitor cells acting at a late stage of megakaryocyte development. It acts as a circulating regulator of platelet numbers.

Inter. Pro: example. . . Examplelist P 33708 P 33709 P 49745 view matches for the examples Publications 1. Shoemaker C. B. , Mitsock L. D. 849 -858 (1986) 2. Takeuchi M. , Takasaki S. , Miyazaki H. , Kato T. , Hoshi S. , Kochibe N. , Kobata A. J. Biol. Chem. 263: 3657 -3663 (1988) 3. Lin F. K. , Lin C. H. , Lai P. H. , Browne J. K. , Egrie J. C. , Smalling R. , Fox G. M. , Chen K. K. , Castro M. , Suggs S. Gene 44: 201 -209 (1986) 4. Nagao M. , Suga H. , Okano M. , Masuda S. , Narita H. , Ikura K. , Sasaki R. Nucleotide sequence of rat erythropoietin. 1171: 99 -102 (1992) Children IPR 003013 Signatures PROSITE PS 00817 EPO_TPO PFAM PF 00758 EPO_TPO Matches Table Graphical

Databases 5: mutation/polymorphism Contain informations on sequence variations that are linked or not to genetic diseases; Mainly human but: OMIA - Online Mendelian Inheritance in Animals General db: OMIM HMGD - Human Gene Mutation db SVD - Sequence variation db HGBASE - Human Genic Bi-Allelic Sequences db db. SNP - Human single nucleotide polymorphism (SNP) db Disease-specific db: most of these databases are either linked to a single gene or to a single disease; p 53 mutation db ADB - Albinism db (Mutations in human genes causing albinism) Asthma and Allergy gene db ….

Mutation/polymorphisms: definitions SNPs: single nucleotide polymorphisms c-SNPs: coding single nucleotide polymorphisms SAPs: single amino-acid polymorphisms (Single Nucleotide Polymorphisms within c. DNA sequences) Missense mutation: -> SAP Nonsense mutation: -> STOP Insertion/deletion of nucleotides -> frameshift… ! Numbering of the mutation depends on the db (aa no 1 is not necessary the initiator Met !)

Mutation/polymorphisms db. SNP consortium http: //snp. cshl. org/ db. SNP at NCBI http: //www. ncbi. nlm. nih. gov/SNP/ Bayer, Roche, IBM, Pfizer, Novartis, Motorola…… Mission: develop up to 300, 000 SNPs distributed evenly throughout the human genome and make the informations related to these SNPs available to the public without intellectual property restrictions. The project started in April 1999 and is anticipated to continue until the end of 2001. Collaboration between the National Human Genome Research Institute and the National Center for Biotechnology Information (NCBI) Mission: central repository for both single base nucleotide subsitutions and short deletion and insertion polymorphisms Aug 24, 2000 , db. SNP has submissions for 803557 SNPs. Chromosome 21 db. SNP http: //csnp. isb-sib. ch/ A joint project between the Division of Medical Genetics of the University of Geneva Medical School and the SIB Mission: comprehensive c. SNP (Single Nucleotide Polymorphisms within c. DNA sequences) database and map of chromosome 21

Mutation/polymorphisms Very heterogeneous format; Generally modest size; There are initiatives to standardize and to unify these databases (SVD - Sequence Variation Database project at EBI: HMut. DB)

Databases 6: proteomics Contain informations obtained by 2 D-PAGE: master images of the gels and description of identified proteins Examples: SWISS-2 DPAGE, ECO 2 DBASE, Maize 2 DPAGE, Sub 2 D, Cyano 2 DBase, etc. Format: composed of image and text files Most 2 D-PAGE databases are “federated” and use SWISS-PROT as a master index There is currently no protein Mass Spectrometry (MS) database (not for long…)

Databases 7: 3 D structure Contain the spatial coordinates of macromolecules whose 3 D structure has been obtained by X-ray or NMR studies Proteins represent more than 90% of available structures (others are DNA, RNA, sugars, virus, complex protein/DNA…) PDB (Protein Data Bank), SCOP (structural classification of proteins (according to the secondary structures)), BMRB (Bio. Mag. Res. Bank; RMN results) Future: Homology-derived 3 D structure db.

PDB Protein Data Bank, managed by RCSB Currently there are ~13’ 000 structures for about 4’ 000 different molecules, but far less protein family ! There also databases that contain data derived from PDB. Examples: HSSP (homologyderived secondary structure of proteins), SWISS 3 DIMAGE (images)… Restriction enzyme

PDB: example HEADER COMPND SOURCE AUTHOR REVDAT JRNL JRNL REMARK REMARK REMARK REMARK ……… LYASE(OXO-ACID) 01 -OCT-91 12 CA 2 CARBONIC ANHYDRASE /II (CARBONATE DEHYDRATASE) (/HCA II) 12 CA 3 2 (E. C. 4. 2. 1. 1) MUTANT WITH VAL 121 REPLACED BY ALA (/V 121 A) 12 CA 4 HUMAN (HOMO SAPIENS) RECOMBINANT PROTEIN 12 CA 5 S. K. NAIR, D. W. CHRISTIANSON 12 CA 6 1 15 -OCT-92 12 CA 0 12 CA 7 AUTH S. K. NAIR, T. L. CALDERONE, D. W. CHRISTIANSON, C. A. FIERKE 12 CA 8 TITL ALTERING THE MOUTH OF A HYDROPHOBIC POCKET. 12 CA 9 TITL 2 STRUCTURE AND KINETICS OF HUMAN CARBONIC ANHYDRASE 12 CA 10 TITL 3 /II$ MUTANTS AT RESIDUE VAL-121 12 CA 11 REF J. BIOL. CHEM. V. 266 17320 1991 12 CA 12 REFN ASTM JBCHA 3 US ISSN 0021 -9258 071 12 CA 13 1 12 CA 14 2 12 CA 15 2 RESOLUTION. 2. 4 ANGSTROMS. 12 CA 16 3 12 CA 17 3 REFINEMENT. 12 CA 18 3 PROGRAM PROLSQ 12 CA 19 3 AUTHORS HENDRICKSON, KONNERT 12 CA 20 3 R VALUE 0. 170 12 CA 21 3 RMSD BOND DISTANCES 0. 011 ANGSTROMS 12 CA 22 3 RMSD BOND ANGLES 1. 3 DEGREES 12 CA 23 4 12 CA 24 4 N-TERMINAL RESIDUES SER 2, HIS 3, HIS 4 AND C-TERMINAL 12 CA 25 4 RESIDUE LYS 260 WERE NOT LOCATED IN THE DENSITY MAPS AND, 12 CA 26 4 THEREFORE, NO COORDINATES ARE INCLUDED FOR THESE RESIDUES. 12 CA 27

PDB (cont. ) SHEET 3 S 10 PHE 66 PHE 70 -1 O ASN 67 N LEU 60 12 CA 68 SHEET 4 S 10 TYR 88 TRP 97 -1 O PHE 93 N VAL 68 12 CA 69 SHEET 5 S 10 ALA 116 ASN 124 -1 O HIS 119 N HIS 94 12 CA 70 SHEET 6 S 10 LEU 141 VAL 150 -1 O LEU 144 N LEU 120 12 CA 71 SHEET 7 S 10 VAL 207 LEU 212 1 O ILE 210 N GLY 145 12 CA 72 SHEET 8 S 10 TYR 191 GLY 196 -1 O TRP 192 N VAL 211 12 CA 73 SHEET 9 S 10 LYS 257 ALA 258 -1 O LYS 257 N THR 193 12 CA 74 SHEET 10 S 10 LYS 39 TYR 40 1 O LYS 39 N ALA 258 12 CA 75 TURN 1 T 1 GLN 28 VAL 31 TYPE VIB (CIS-PRO 30) 12 CA 76 TURN 2 T 2 GLY 81 LEU 84 TYPE II(PRIME) (GLY 82) 12 CA 77 TURN 3 T 3 ALA 134 GLN 137 TYPE I (GLN 136) 12 CA 78 TURN 4 T 4 GLN 137 GLY 140 TYPE I (ASP 139) 12 CA 79 TURN 5 THR 200 LEU 203 TYPE VIA (CIS-PRO 202) 12 CA 80 TURN 6 T 6 GLY 233 GLU 236 TYPE II (GLY 235) 12 CA 81 CRYST 1 42. 700 41. 700 73. 000 90. 00 104. 60 90. 00 P 21 2 12 CA 82 ORIGX 1 1. 000000 0. 00000 12 CA 83 ORIGX 2 0. 000000 1. 000000 0. 00000 12 CA 84 ORIGX 3 0. 000000 1. 000000 0. 00000 12 CA 85 SCALE 1 0. 023419 0. 000000 0. 006100 0. 00000 12 CA 86 SCALE 2 0. 000000 0. 023981 0. 000000 0. 00000 12 CA 87 SCALE 3 0. 000000 0. 014156 0. 00000 12 CA 88 ATOM 1 N TRP 5 8. 519 -0. 751 10. 738 1. 00 13. 37 12 CA 89 ATOM 2 CA TRP 5 7. 743 -1. 668 11. 585 1. 00 13. 42 12 CA 90 ATOM 3 C TRP 5 6. 786 -2. 502 10. 667 1. 00 13. 47 12 CA 91 ATOM 4 O TRP 5 6. 422 -2. 085 9. 607 1. 00 13. 57 12 CA 92 ATOM 5 CB TRP 5 6. 997 -0. 917 12. 645 1. 00 13. 34 12 CA 93 ATOM 6 CG TRP 5 5. 784 -0. 209 12. 221 1. 00 13. 40 12 CA 94 ATOM 7 CD 1 TRP 5 5. 681 1. 084 11. 797 1. 00 13. 29 12 CA 95 ATOM 8 CD 2 TRP 5 4. 417 -0. 667 12. 221 1. 00 13. 34 12 CA 96 ATOM 9 NE 1 TRP 5 4. 388 1. 418 11. 515 1. 00 13. 30 12 CA 97 ATOM 10 CE 2 TRP 5 3. 588 0. 375 11. 797 1. 00 13. 35 12 CA 98 ATOM 11 CE 3 TRP 5 3. 837 -1. 877 12. 645 1. 00 13. 39 12 CA 99 ATOM 12 CZ 2 TRP 5 2. 216 0. 208 11. 656 1. 00 13. 39 12 CA 100 ATOM 13 CZ 3 TRP 5 2. 465 -2. 043 12. 504 1. 00 13. 33 12 CA 101 ATOM 14 CH 2 TRP 5 1. 654 -1. 001 12. 009 1. 00 13. 34 12 CA 102 …….

Databases 8: metabolic Contain informations that describe enzymes, biochemical reactions and metabolic pathways; ENZYME and BRENDA: nomenclature databases that store informations on enzyme names and reactions; Examples of metabolic databases: Eco. Cyc (specialized on Escherichia coli), KEGG, EMP/WIT; Usualy these databases are tightly coupled with query software that allows the user to visualise reaction schemes.

Databases 9: bibliographic Bibliographic reference databases contain citations and abstract informations of published life science articles; Example: Medline Other more specialized databases also exist (example: Agricola).

Medline MEDLINE covers the fields of medicine, nursing, dentistry, veterinary medicine, the health care system, and the preclinical sciences more than 4, 000 biomedical journals published in the United States and 70 other countries Contains over 10 million citations since 1966 until now Contains links to biological db and to some journals New records are added to Pre. MEDLINE daily! Many papers not dealing with human are not in Medline ! Before 1970, keeps only the first 10 authors ! Not all journals have citations since 1966 !

Medline/Pubmed Pub. Med is developed by the National Center for Biotechnology Information (NCBI) Pub. Med provides access to bibliographic information such as MEDLINE, Pre. MEDLINE, Health. STAR, and to integrated molecular biology databases (composite db) PMID: 10923642 (Pub. Med ID), UI: 20378145 (Medline ID)

Databases 10: others There are many databases that cannot be classified in the categories listed previously; Examples: Re. Base (restriction enzymes), TRANSFAC (transcription factors), OGLYCBASE (O-linked sugars), Protein-protein interactions db (DIR), biotechnology patents db, etc. ; As well as many other resources concerning any aspects of macromolecules and molecular biology.

Proliferation of databases What is the best db for sequence analysis ? Which does contain the highest quality data ? Which is the more comprehensive ? Which is the more up-to-date ? Which is the less redundant ? Which is the more indexed (allows complex queries) ? Which Web server does respond most quickly ? ……. ? ? ?

Some important practical remarks Databases: many errors (automated annotation) ! Not all db are available on all servers The update frequency is not the same for all servers; creation of db_new between releases (exemple: EMBLnew; Tr. EMBLnew…. ) Some servers add automatically useful cross-references to an entry (implicit links) in addition to already existing links (explicit links)

Database retrieval tools Sequence Retrieval System (SRS, Europe) allows any flat-file db to be indexed to any other; allows to formulate queries across a wide range of different db types via a single interface, without any worry about data structure, query languages… Entrez (USA): less flexible than SRS but exploits the concept of « neighbouring » , which allows related articles in different db to be linked together, whether or not they are cross-referenced directly ATLAS: specific for macromolecular sequences db (i. e. NRL-3 D) ….

More informations about SWISSPROT

The golden goals of SWISS-PROT Annotated / curated Complete Non-redundant Highly cross-referenced Available from a variety of servers and through sequence analysis software tools Associated with wide range of documentation Review: Protein sequence databases R. Apweiler (2000), Adv. in protein chemistry, 54, 31 -70

SWISS-PROT: species 6’ 840 different species 20 species represent about 45% of all sequences in the database 5’ 000 species are only represented by one to three sequences. In most cases, these are sequences which were obtained in the context of a phylogenetic study

SWISS-PROT: cross-references SWISS-PROT was the first database with crossreferences. Explicitly cross-referenced to 34 databases Cross-ref to DNA (EMBL/Gen. Bank/DDBJ), 3 Dstructure (PDB), literature (Medline), genomic (MIM, MGD, Fly. Base, SGD, Subti. List, etc. ), 2 D-gel (SWISS-2 DPAGE), specialized db (PROSITE, TRANSFAC) Implicitly cross-referenced to additional db on the WWW (Gene. Cards, PRODOM, etc. )

Annotations Function(s) Post-translational modifications (PTM) Domains Quaternary structure Similarities Diseases, mutagenesis Conflicts, variants Cross-references …

A Swiss-Prot entry

Sprot entry (cont. )

Future for human proteins Original estimate: from 70’ 000 to 100’ 000 genes Incyte recently announced an estimation of 140’ 000 genes More recent estimations give about 30’ 000 to 40’ 000 genes C. elegans and Drosophila have ~15’ 000 genes. There was two sets of genome duplication in the evolutionary history leading to vertebrates. Very roughly it means that: Human genes=~60’ 000 genes - losses + new genes But more than 1 million proteins ! (due to PTM, alternative products, variants…) http: //www. ensembl. org/genesweep. html

Genesweep http: //www. ensembl. org/genesweep. html

What after genomes? Proteome projects are an essential tool for the understanding of real proteins There will be a flood of characterization data (MS, 2 D) that will be the equivalent of ESTs at the protein level Protein databases are going to be more and more important for new biological studies

Databases in GCG DNA Protein EMBL, EPD, Rep. Base, vectordb (NCBI) Swiss-Prot, Tr. EMBL, PDB Other PROSITE, REBASE

How to access databases in GCG? Fetch or typedata ? Stringsearch Name Lookup (based on SRS) Useful to generate list files