Shared Active Site Architecture between the Large Subunit of Eukaryotic Primase and DNA Photolyase 英文参考文献.docVIP

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Shared Active Site Architecture between the Large Subunit of Eukaryotic Primase and DNA Photolyase 英文参考文献.doc

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Shared Active Site Architecture between the Large Subunit of Eukaryotic Primase and DNA Photolyase 英文参考文献

SharedActiveSiteArchitecturebetweentheLarge SubunitofEukaryoticPrimaseandDNAPhotolyase LudovicSauguet,SebastianKlinge¤,RajikaL.Perera,JosephD.Maman,LucaPellegrini* DepartmentofBiochemistry,UniversityofCambridge,Cambridge,UnitedKingdom Abstract Background: DNA synthesis during replication relies on RNA primers synthesised by the primase, a specialised DNA- dependent RNA polymerase that can initiate nucleic acid synthesis de novo. In archaeal and eukaryotic organisms, the primaseisaheterodimericenzymeresultingfromtheconstitutiveassociationofasmall(PriS)andlarge(PriL)subunit.The abilityoftheprimasetoinitiatesynthesisofanRNAprimerdependsonaconservedFe-SdomainattheC-terminusofPriL (PriL-CTD).However,thecriticalroleofthePriL-CTDinthecatalyticmechanismofinitiationisnotunderstood. Methodology/Principal Findings: Here we report the crystal structure of the yeast PriL-CTD at 1.55A? resolution. The structurerevealsthatthePriL-CTDfoldsintwolargelyindependentalpha-helicaldomainsjoinedattheirinterfacebya[4Fe- 4S]cluster.ThelargerN-terminaldomainrepresentsthemostconservedportionofthePriL-CTD,whereasthesmallerC- terminal domain is largely absent in archaeal PriL. Unexpectedly, the N-terminal domain reveals a striking structural similaritywiththeactivesiteregionoftheDNAphotolyase/cryptochromefamilyofflavoproteins.Theregionofsimilarity includesPriL-CTDresiduesthatareknowntobeessentialforinitiationofRNAprimersynthesisbytheprimase. Conclusion/Significance:OurstudyreportsthefirstcrystallographicmodeloftheconservedFe-Sdomainofthearchaeal/ eukaryotic primase. The structural comparison with a cryptochrome protein bound to flavin adenine dinucleotide and single-strandedDNAprovidesimportantinsightintothemechanismofRNAprimersynthesisbytheprimase. Citation:SauguetL,KlingeS,PereraRL,MamanJD,PellegriniL(2010)SharedActiveSiteArchitecturebetweentheLargeSubunitofEukaryoticPrimaseandDNA Photolyase.PLoSONE5(4):e10083.doi:10.1371/journal.pone.0010083 Editor:BostjanKobe,UniversityofQueensland,Austral