In vitro recombination catalyzed by bacterial class 1 integron integrase IntI1 involves cooperative binding and specific oligomeric intermediates.
Gene transfer via bacterial integrons is a major pathway for facilitating the spread of antibiotic resistance genes across bacteria. Recently the mechanism underlying the recombination catalyzed by class 1 integron recombinase (IntI1) between attC and attI1 was highlighted demonstrating the involvement of a single-stranded intermediary on the attC site. However, the process allowing the generation of this single-stranded substrate has not been determined, nor have the active IntI1*DNA complexes been identified. Using the in vitro strand transfer assay and a crosslink strategy we previously described we demonstrated that the single-stranded attC sequences could be generated in the absence of other bacterial proteins in addition to IntI. This suggests a possible role for this protein in stabilizing and/or generating this structure. The mechanism of folding of the active IntI*DNA complexes was further analyzed and we show here that it involves a cooperative binding of the protein to each recombination site and the emergence of different oligomeric species specific for each DNA substrate. These findings provide further insight into the recombination reaction catalyzed by IntI1.
Dubois
Véronique
V
Laboratory of Cellular and Molecular Microbiology and Pathogenicity (MCMP), UMR 5097-CNRS, University Victor Segalen Bordeaux 2, Bordeaux, France.
Debreyer
Carole
C
Quentin
Claudine
C
Parissi
Vincent
V
D001419:Bacteria / Q000201:enzymology*; D001483:Base Sequence; D055162:Biocatalysis; D001704:Biopolymers / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D019426:Integrases / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides; D011485:Protein Binding; D011995:Recombination, Genetic
D001704:Biopolymers; D009838:Oligodeoxyribonucleotides; D019426:Integrases; C115006:integron integrase IntI1