A target specificity switch in IS911 transposition: the role of the OrfA protein.
The role played by insertion sequence IS911 proteins, OrfA and OrfAB, in the choice of a target for insertion was studied. IS911 transposition occurs in several steps: synapsis of the two transposon ends (IRR and IRL); formation of a figure-of-eight intermediate where both ends are joined by a single-strand bridge; resolution into a circular form carrying an IRR-IRL junction; and insertion into a DNA target. In vivo, with OrfAB alone, an IS911-based transposon integrated with high probability next to an IS911 end located on the target plasmid. OrfA greatly reduced the proportion of these events. This was confirmed in vitro using a transposon with a preformed IRR-IRL junction to examine the final insertion step. Addition of OrfA resulted in a large increase in insertion frequency and greatly increased the proportion of non-targeted insertions. The intermolecular reaction leading to targeted insertion may resemble the intramolecular reaction involving figure-of-eight molecules, which leads to the formation of circles. OrfA could, therefore, be considered as a molecular switch modulating the site-specific recombination activity of OrfAB and facilitating dispersion of the insertion sequence (IS) to 'non-homologous' target sites.
Loot
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Laboratoire de Microbiologie et Génétique Moléculaire, CNRS UMR5100, 118 Rte de Narbonne, F-31062 Toulouse Cedex, France.
Turlan
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C
Rousseau
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Ton-Hoang
B
B
Chandler
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M
D001704:Biopolymers; D004251:DNA Transposable Elements / Q000235:genetics*; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D004270:DNA, Circular / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics / Q000502:physiology*; D015964:Gene Expression Regulation, Bacterial; D046911:Macromolecular Substances; D008957:Models, Genetic; D016254:Mutagenesis, Insertional; D011995:Recombination, Genetic; D013379:Substrate Specificity; D019895:Transposases / Q000235:genetics / Q000502:physiology*
D001704:Biopolymers; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D004270:DNA, Circular; D029968:Escherichia coli Proteins; D046911:Macromolecular Substances; C085896:OrfB protein, E coli; D019895:Transposases