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PMID: 2970060 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

An intermediate in the phage lambda site-specific recombination reaction is revealed by phosphorothioate substitution in DNA.

Nucleic acids research ·Vol. 16 ·No. 14B ·1988-07-25 ·Pages 6839-56

Kitts PA, Nash HA

Abstract

It has been proposed that phage lambda site-specific recombination proceeds via two independent strand exchanges: the first exchange forming a Holliday-structure which is then converted into complete recombinant products by the second strand exchange. If this hypothesis is correct, one should be able to trap the putative Holliday intermediate by preventing the second strand exchange. In this paper, we show that substitution of phosphorothioate for phosphate in one strand of a recombination site is an effective way to block recombination while permitting the accumulation of a novel structure. This effect is seen only when phosphorothioate is positioned at a point of potential cleavage by Int recombinase, demonstrating that the inhibition of strand exchange is highly specific. Analysis of the novel structure that accumulates in these reactions proves that it contains a Holliday joint. Holliday-structures can also be detected in unblocked recombinations but are present at very low levels. The characteristics of Holliday-structure formation that we describe substantiate the proposed recombination pathway.

MeSH Terms
Bacteriophage lambda/genetics Chemical Phenomena Chemistry DNA, Bacterial/genetics Escherichia coli/genetics In Vitro Techniques Organothiophosphorus Compounds Phosphates/metabolism Recombination, Genetic
Chemicals
DNA, Bacterial Organothiophosphorus Compounds Phosphates
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Kitts P A
Laboratory of Molecular Biology, National Institute of Mental Health, Bethesda, MD 20892.
Nash H A
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Article Info
Journal
Nucleic acids research
Abbr.
Nucleic Acids Res
ISSN
0305-1048
DOI
10.1093/nar/16.14.6839
Published
1988-07-25
Pages
6839-56
Language
English
Region
England
NLM ID
0411011
PMCID
PMC338337
Subset
IM
External Links
PubMed source DOI: 10.1093/nar/16.14.6839 PMC Full Text View journal details
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