Protein Domain Identification Curso Doctorado UAM 2008
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1 ProteinDomain Identification CursoDoctorado UAM 2008 LuisSánchezPulido CentroNacionaldeBiotecnología Madrid
2 Whydoweanalysesequences? Proteinswithknownsequence 3D Function Both?????
3 DataOverload!!! Databasegrowthbyyear www3.ebi.ac.uk/services/dbstats/
4 Thankstotherecognitionof homologybetweenproteins, wecan TRANSFERINFORMATION Structuraland/orFunctional
5 Homologues:twoproteinswithacommonancestor....dependentonthetypeofdivergencetheycanbe: orthologues speciation paralogues geneduplication xenologues horizontaltransference
6 INFORMATIONTRANSFER Structural fromhomologousproteinsofknownstructure(x Ray,NMRoEM) Functional fromexperimentallycharacterisedhomologousproteinsortheirgenomic orproteomiccontext
7 TheStructureisbetterconservedthansequence! D'AlfonsoG,TramontanoA,LahmA. Structuralconservationinsingle domainproteins:implicationsforhomologymodeling. JStructBiol.134, (2001)
8 ARemoteHomologyexample: 1NYN 1P9Q SBDSFamily
9 DefiniendoHomologíaRemota truepositives RostB.(1999) Twilightzoneof proteinsequencealignments. ProteinEng.12: truenegatives
10 Comparisonsbetweenpairsofsequenceswithknownstructure 100 Identity 50 20% Size Twilight zone Chothia&Lesk, 1986 Rost,1999 == Rmsd>3ARmsd<3A
11 INFORMATIONTRANSFER Structural fromhomologousproteinsofknownstructure(x Ray,NMRoEM) Functional fromexperimentallycharacterisedhomologousproteinsortheirgenomic orproteomiccontext
12 FUNCTION? Thesearehomologous Proteins... Theirroleinthecellis verydifferent But...AllofthembindGTP
13 Depending onthe Definition offunction
14 TheTransferofStructuraland/orFunctional Informationbetweenhomologousproteinsisa ComplexTask Howisitdone? DivideeachoftheTasksin asmanypartsasisnecessary toresolvetheproblem
15 Domain Definition Domains are described, from a structural point of view, as structurally compact units, locally independent in function and folding and usually characterized by a well define hydrophobic core. From sequence analysis point of view, we describe domains as evolutionary conserved regions that are present in different protein families of diverse architecture. Hypothetical Domain
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17 REPEATS InthelimitsofDomainDefinition LRR HEAT TPR PFTA beta l WD40 Proteinrepeats.ShortspecialistreviewfortheEncyclopediaofGenomics,Proteomics,and Cedidapor:Perez IratxetaC,AndradeMA(2005)Bioinf.Ed.WileyandSonsLtd.,UK.
18 Proteinirregularitiesthathindersequence analysis Lowcomplexityregions Repeats,Trans membraneandcoiled coilregions(highmutation rates) andfoldirregularities,suchas: CircularPermutationsandInsertions
19 Theroleofdomainsinprotein evolution Shuffling,AccretionandSupra Domains VERSATILITY!!
20 METHODS ON DOMAIN ORIENTED SEQUENCE ANALYSIS
21 CommonName IDorACCorGI ReferenceDatabase SRS - EBI Entrez NCBI Buscar comparando: Sequence MRTSRGH... Alignment >Profile RTNMSDAQQ GSWYSDPK REGWFYN RTNMSDAQQ GSWYSDPK REGWFYN RTNMSDAQQ GSWYSDPK REGWFYN RTNMSDAQQ GSWYSDPK REGWFYN RTNMSDAQQ GSWYSDPK REGWFYN RTNMSDAQQ GSWYSDPK REGWFYN RTNMSDAQQ GSWYSDPK REGWFYN Secuencia contra Secuencias BLAST Secuencias contra Perfiles Pfam Buscar comparando: Perfil contra Secuencias PsiBlast o HMMer Perfil contra Perfiles HHpred
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30 Detectionofhomologousproteinsequences Profiles (PSSMs:PsiBlast,HMMs) Reciprocal!
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38 TwomainCharacterictics: CombiningMultipleAlignmentMethods MixingHeterogenousInformation AND
39 Combining Multiple Alignment Methods Clustal W T-Coffee Probcons Specialist Muscle T-Coffee Multiple Sequence Alignment CopyrightCédricNotredame,2000,allrightsreserved
40 Mixing Heterogenous Information Local Alignment Global Alignment Multiple Alignment Specialist Structural T-Coffee Multiple Sequence Alignment CopyrightCédricNotredame,2000,allrightsreserved
41 Duringthistediouswork... ofsequencebysequencecompilation Alotofblastsearchestowarddifferent(ests,genomicDNAandProtein)sequencedatabases ESTclusteringandtranscriptreconstruction IncludingGeneFinding(FGENESH+)... WemusthaveFaithinthat: Everynewsequencethatweaddtothealignment, couldbecausefor"phasetransition"between aprofilethatlocalize,ornot,remotehomologousproteins...withknownfunction!!!
42 Admiring the amazing life's diversity GenBank CopyrightCédricNotredame,2000,allrightsreserved dbest
43 BasicReferences: ZuckerkandlE,yPaulingL.(1965) Evolutionarydivergenceandconvergenceinproteins. EvolvingGenesandProteins, AcademicPress,NewYork, Questions:
44 CommonName IDorACCorGI BasedeDatosdeReferencia SRS - EBI Entrez NCBI UNIPROT Buscar comparando: Sequence MRTSRGH... Alignment >Profile RTNMSDAQQ GSWYSDPK REGWFYN RTNMSDAQQ GSWYSDPK REGWFYN RTNMSDAQQ GSWYSDPK REGWFYN RTNMSDAQQ GSWYSDPK REGWFYN RTNMSDAQQ GSWYSDPK REGWFYN RTNMSDAQQ GSWYSDPK REGWFYN RTNMSDAQQ GSWYSDPK REGWFYN Secuencia contra Secuencias BLAST Secuencias contra Perfiles Pfam Buscar comparando: Perfil contra Secuencias PsiBlast o HMMer Perfil contra Perfiles HHpred
45 Detectionofhomologousproteinsequences Profiles (PSSMs:PsiBlast,HMMs) Reciprocal!
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47 A easier way to view a Protein Sequence... Thanks to DOMAIN DataBases
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50 Pfam IgCLANFamily
51 Pfam IgCLANFamily
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57 STRINGUPDATE(Version7.0) available1stjanuary2007 WhatisSTRING?? MoreGenomes 0.7millionsofproteinsin179species 1.5millionsofproteinsin373species CoreyPeriphera l STRING&SMART MoreDatabases... MINT,HPRD,BioGRID,DIPandReactome NewDisplay AJAX
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65 Sequence Domain Oriented Sequence Sequence DataBases dbest Analysis Flow-Chart HMMer GenBank Domain Databases As you will never be sure which are the right problems to work on, most of the time that you spend in the laboratory or at your desk will be wasted. If you want to be creative, HHpred ALIGNMENT HMMer then you will have to get used to spending most of your time DOMAIN Hypothetical not being creative, Domain to being becalmed on the ocean of scientific knowledge. Steven Weinberg Biochemical Knowledge How Do the Pieces of the Functional Assignment Puzzle Fit Together? Functional Hypothesis
66 REAL-LIFE EXAMPLES
67 SPOC:Awidelydistributeddomainassociatedwithcancer,apoptosisandtranscription. Sanchez PulidoL,RojasAM,VanWelyK,Martinez AC,ValenciaA. CNB CSIC DATF1_HUMAN(DIDO 1) TFS2M PHD dphd SPOC
68 Domainlocatedin, atleast, TwoArchitectures
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70 Gas1isrelatedtotheGFRαfamilyandregulatesRetsignaling CabreraJ.R.,Sánchez PulidoL.,RojasA.M.,ValenciaA., MańesS.,NaranjoJ.R.&MellstromB.(2005) CNB CSIC InitialProtein:GAS1(GrowthArrestSpecific1) FUNCIÓN:Regulacióndeprocesosapoptóticos. Enunprimerabordaje: Péptidoseńal DuplicaciónInterna GPI Anclaje
71 GAS1&GFR α ModelStructure
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73 Arquitecturasimilar
74 JRCabrera CNB CSIC BIBLIOGRAFÍA Diferentesfactorestróficosoligandos: GDNFGlialcellDerivedNeurotrophicFactor NRTNNeurturin ARTNArtemin PSPNPersephin LaFamiliaGFRαconverge enlatransduccióndelaseńal ensuinteraccióncon laquinasaret > YGAS1...?
75 Computational predictions supportedby experimentalanalysis.
76 DTRGHYFASSTNDR???????????? JorgeRuben, Santos & Ana
77 MALproteinFamily MARVEL:aconserveddomaininvolvedinmembraneappositionevents. Sánchez PulidoL.,Martín BelmonteF.,ValenciaA.&AlonsoM.A.(2002) CNB CSIC&CBM CSIC InitialProtein:MAL Function:MembraneTraffic ApicalZoneofpolarizedEpithelialCells TransGolgi Endosomes Membrane
78 Diverse...buthasCOMMONFunctionalElements Rafts=Cholesterol+Sphingolipids Zonulaoccludens CONCLUSION: Similar MARVELdomaincouldbeusedasmachineryforraft Phenotypesin organizationinmembraneappositionevents,suchas ChristophThiele,MatthewJ.Hannah,Falk thoseoccurringduringbiogenesisoftransportvesicles Fahrenholz&WielandB.Huttner OverExpression (e.g.mal,physinsandgyrins)ortheformationof NatureCellBiology2,(2000) Cholesterolbindstosynaptophysinandis specializedclosecontacts(kisses)intightjunctions requiredforbiogenesisofsynapticvesicles (e.g.occludin).
79 ?????????????????? REL RER Endocitosis Lisosomas Secrección Vesícula Golgi HELPinexperimentalCharacterization
80 p40 rtt45 Tinton23 p23 Rr78 SinChan2 ppr..... EssentialGenesinMyoblatfusion MARVEL DTRGHYFASSTNDR???????????? HELPinexperimentalCharacterization
81 CONCLUSION Cada Proteína es un MUNDO! Asageneralguidetofunctionalannotation,itshouldbekept inmindthatcurrentmethodsforgenomeanalysis,eventhe mostpowerfulandsophisticatedofthem,facilitate,butdo notsupplanttheworkofahumanexpert. EugeneKoonin.
82 Ofallnaturalsystems,livingmatter istheonewhich,inthefaceofgreattransformations, preservesinscribedinitsorganizationthelargestamountofitsownpasthistory. UsingHegel'sexpression,wemaysaythatthereisnoothersystemthatisbetter"aufgehoben" (constantlyabolishedandsimultaneouslypreserved). Wemayaskthequestionswhereinthenowlivingsystemsthegreatestamount oftheirpasthistoryhassurvivedand Bioinformatics howitcanbeextracted. EmileZuckerkandlyLinusPauling,1965. Moleculesasdocumentsofevolutionaryhistory. JTheorBiol.8: Evolutionarydivergenceandconvergenceinproteins. EvolvingGenesandProteins,AcademicPress,NewYork,
83 BasicReferences: ZuckerkandlE,yPaulingL.(1965) Evolutionarydivergenceandconvergenceinproteins. EvolvingGenesandProteins, AcademicPress,NewYork, BorkP,GibsonTJ.(1996) Applyingmotifandprofilesearches. MethodsEnzymol.266: IyerLM,AravindL,BorkP,HofmannK,MushegianAR,ZhulinIB,KooninEV.(2001) Quoderatdemonstrandum?Themysteryofexperimentalvalidationofapparentlyerroneous computationalanalysesofproteinsequences. GenomeBiol.2:RESEARCH0051. Questions:
Linear Sequence Analysis. 3-D Structure Analysis
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