Chapter 22 Homologous Recombination @Rafferty,J.B.,S.E.Sedelnikova,D.Hargreaves,P.J.Artymink,P.J.Baker,G.J.Sharples, A.A.Mahdi,R.G.Lloyd,and D.W.Rice,Crystal structure of DNA recombination protein RuvA
Chapter 22 Homologous Recombination
homologous recombination:a crossover()event that joins DNA segments that were previously separated.Recombination can be intramolecular(分子内的)or intrermolecular. CopyrightThe McGraw-Hill Companies,Inc Permission required for reproduction or display Intermolecular: (a)Single crossover: (b)Double crossover Intramolecular: (a)Direct repeats: (b)Inverted repeats:
homologous recombination: a crossover(交换)event that joins DNA segments that were previously separated．Recombination can be intramolecular(分子内的) or intrermolecular
22.1 The RecBCD Pathway for Homologous Recombination(同源重组的RecBCD途径) The recombination process begins when the RecBCD protein,the product of the recB,-C and-D genes, binds to a DNA double-stranded break(DSB)and uses its DNA helicase activity to unwind the DNA toward a so-called chi site,x,which has the sequence 5'- GCTGGTGG-3'.Chi sites are found on average every 5000 bp in the E.coli genome. RecBCD also has double-stranded and single- stranded exonuclease and single-stranded endonuclease activities.These allow RecBCD to produce a single-stranded tail,which can then be coated by RecA protein
22.1 The RecBCD Pathway for Homologous Recombination(同源重组的RecBCD途径) ✓ The recombination process begins when the RecBCD protein，the product of the recB，-C and-D genes， binds to a DNA double-stranded break(DSB) and uses its DNA helicase activity to unwind the DNA toward a so-called chi site,χ，which has the sequence 5’- GCTGGTGG-3’. Chi sites are found on average every 5000 bp in the E.coli genome． ✓ RecBCD also has double-stranded and singlestranded exonuclease and single-stranded endonuclease activities．These allow RecBCD to produce a single-stranded tail，which can then be coated by RecA protein．
5 ◆3 Chi a protruding end.cc (a)The RecBCD proteinbinds at a double- stranded DNA break,and the DNA helicase (b)Strand invasion:D-loop formation activity of RecBCD then unwinds the DNA cc toward a Chi site,ultimately creating a 3'- Scan for homology. ss DNA that is coated with RecA protein and SSB. (b)RecA promotes invasion of another DNA dy Nick duplex,forming a D loop. Strand exchange (RecA+SSB) (C)RecA helps the invading strand scan for a region of homology in the recipient DNA (e) Repair gaps and seal nicks (Holliday junction) duplex.Here,the invading strand has base- 3 paired with a homologous region,releasing Branch migration(RuvA RuvB) SSB and RecA. (g)Resolution (RuvC) ative resolution (RuvC)
(a)The RecBCD proteinbinds at a doublestranded DNA break，and the DNA helicase activity of RecBCD then unwinds the DNA toward a Chi site，ultimately creating a 3’- ss DNA that is coated with RecA protein and SSB. (b)RecA promotes invasion of another DNA duplex, forming a D loop． (C)RecA helps the invading strand scan for a region of homology in the recipient DNA duplex．Here, the invading strand has basepaired with a homologous region，releasing SSB and RecA
opyright The McGraw-Hl Compar ■5 3 (d)Once a homologous region is found,a nick Chi in the looped-out DNA appears,perhaps (a)RecBCD unwinds DNA and leaves 3'-protruding end,coated with RecA caused by RecBCD.This permits the tail of 0G00000003 the newly nicked DNA to base-pair with the (b) Strand invasion:D-loop formation single-stranded region in the other DNA, probably aided by RecA and SSB. (c) Scan for homology (e)The remaining gap are filled in and nicks are sealed by DNA ligase.yielding a four stranded complex with a Holliday junction. Nick Strand exchange(RecA+SSB) (f)Branch migration occurs,sponsored by RuvA and RuvB.Notice that the branch has Repair gaps and seal nicks migrated to the right. (Holliday junction) (g and h)Nicking by RuvC resolves the 3 ◆3 structure into two molecules,crossover 15 Branch migration (RuvA+RuvB) recombinants or heteroduplexesheteroduplex (异源双链核酸分子). (g)Resolution(RuvC)
(d)Once a homologous region is found，a nick in the looped-out DNA appears, perhaps caused by RecBCD．This permits the tail of the newly nicked DNA to base-pair with the single-stranded region in the other DNA， probably aided by RecA and SSB． (e)The remaining gap are filled in and nicks are sealed by DNA ligase．yielding a four stranded complex with a Holliday junction． (f) Branch migration occurs, sponsored by RuvA and RuvB．Notice that the branch has migrated to the right． (g and h) Nicking by RuvC resolves the structure into two molecules，crossover recombinants or heteroduplexesheteroduplex (异源双链核酸分子)．