Isolation and Characterization of Intermediates in Site-Specific Recombination

Cre, the site-specific recombinase from bacteriophage P1, catalyzes a recombination reaction between specific DNA sequences designated as lox sites. The breakage and rejoining of partners during this recombination process must be highly concerted because it has not been possible to detect intermedia...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 1987-10, Vol.84 (19), p.6840-6844
Main Authors: Hoess, Ronald, Wierzbicki, Anna, Abremski, Kenneth
Format: Article
Language:English
Subjects:
Bacteriophages
Biological and medical sciences
Coliphages - genetics
Cruciform DNA
DNA
DNA Nucleotidyltransferases - genetics
DNA Nucleotidyltransferases - metabolism
DNA Restriction Enzymes
DNA, Viral - genetics
DNA, Viral - ultrastructure
Electron microscopy
Enzymes
Escherichia coli - genetics
Fundamental and applied biological sciences. Psychology
Gels
Genic rearrangement. Recombination. Transposable element
Integrases
Molecular and cellular biology
Molecular genetics
Molecules
Mutation
Plasmids
Recombination reactions
Recombination, Genetic
Topology
Viral Proteins
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Summary:Cre, the site-specific recombinase from bacteriophage P1, catalyzes a recombination reaction between specific DNA sequences designated as lox sites. The breakage and rejoining of partners during this recombination process must be highly concerted because it has not been possible to detect intermediates of the reaction with wild-type Cre. Several mutant Cre proteins have been isolated that produce significant amounts of a possible intermediate product of the recombination reaction. The product has been identified as a Holliday structure in which one set of the DNA strands of the recombining partners has been exchanged. Wild-type Cre protein is capable of acting on this structure to form recombinant products, which is consistent with this being an intermediate in the recombination reaction. Characterization of the Holliday structure indicated that one set of strands in the recombining partners was always exchanged preferentially before the other set. In addition, it has been found that certain Cre mutants that are unable to carry out recombination in vitro are able to resolve the intermediate. This suggests that these mutants are defective in a step in the reaction that precedes the formation of the Holliday intermediate.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.84.19.6840