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Partial reconstitution of DNA large loop repair with purified proteins from Saccharomyces cerevisiae

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dc.contributor.author Lahue, Robert S. en
dc.date.accessioned 2009-12-14T09:46:36Z en
dc.date.available 2009-12-14T09:46:36Z en
dc.date.issued 2008 en
dc.identifier.citation Sommer D., Stith C. M., Burgers P. M. J. and Lahue R. S. (2008). Partial reconstitution of DNA large loop repair with purified proteins from Saccharomyces cerevisiae. "Nucleic Acids Research" 36(14) 4699-4707. en
dc.identifier.uri http://hdl.handle.net/10379/515 en
dc.description.abstract Small looped mispairs are corrected by DNA mismatch repair. In addition, a distinct process called large loop repair (LLR) corrects heteroduplexes up to several hundred nucleotides in bacteria, yeast and human cells, and in cell-free extracts. Only some LLR protein components are known, however. Previous studies with neutralizing antibodies suggested a role for yeast DNA polymerase (Pol ), RFC and PCNA in LLR repair synthesis. In the current study, biochemical fractionation studies identified FEN1 (Rad27) as another required LLR component. In the presence of purified FEN1, Pol , RFC and PCNA, repair occurred on heteroduplexes with loops ranging from 8 to 216 nt. Repair utilized a 5' nick, with correction directed to the nicked strand, irrespective of which strand contained the loop. In contrast, repair of a G/T mismatch occurred at low levels, suggesting specificity of the reconstituted system for looped mispairs. The presence of RPA enhanced reactivity on some looped substrates, but RPA was not required for activity. Although additional LLR factors remain to be identified, the excision and resynthesis steps of LLR from a 5' nick can be reconstituted in a purified system with FEN1 and Pol , together with PCNA and its loader RFC. en
dc.format application/pdf en
dc.language.iso en en
dc.publisher OUP en
dc.subject.lcsh Saccharomyces cerevisiae en
dc.subject.lcsh DNA repair en
dc.title Partial reconstitution of DNA large loop repair with purified proteins from Saccharomyces cerevisiae en
dc.type Article en
dc.local.publishedsource http://dx.doi.org/10.1093/nar/gkn446 en
dc.description.peer-reviewed peer-reviewed en

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