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DNA structure specificity of Rap endonuclease.

Sharples, G.J.; Corbett, L.M.; McGlynn, P.; Bolt, E.L.


L.M. Corbett

P. McGlynn

E.L. Bolt


The Rap protein of phage lambda is an endonuclease that nicks branched DNA structures. It has been proposed that Rap can nick D-loops formed during phage recombination to generate splice products without the need for the formation of a 4-strand (Holliday) junction. The structure specificity of Rap was investigated using a variety of branched DNA molecules made by annealing partially complementary oligo-nucleotides. On Holliday junctions, Rap endonuclease shows a requirement for magnesium or manganese ions, with Mn(2+)supporting 5-fold more cleavage than Mg(2+). The location of endonuclease incisions was determined on 3'-tailed D-loop, bubble, flayed duplex, 5'-flap and Y junction DNA substrates. In all cases, Rap preferentially cleaves at the branch point of these molecules. With a flayed duplex, incisions are made in the duplex adjacent to the single-strand arms. Comparison of binding and cleavage specificities revealed that Rap is highly structure-specific and exhibits a clear preference for 4- and 3-stranded DNA over Y and flayed duplex DNA. Almost no binding or cleavage was detected with duplex, partial duplex and single-stranded DNA. Thus Rap endonuclease shows a bias for structures that resemble D-loop and Holliday junction recombination intermediates.


Sharples, G., Corbett, L., McGlynn, P., & Bolt, E. (1999). DNA structure specificity of Rap endonuclease. Nucleic Acids Research, 27, 4121-4127

Journal Article Type Article
Publication Date 1999
Journal Nucleic Acids Research
Print ISSN 0305-1048
Publisher Oxford University Press
Volume 27
Pages 4121-4127
Publisher URL