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Papers In Press, published online ahead of print August 10, 2000
Biology, San Diego State University, San Diego, CA 92182-4614
Corresponding Author: asegall{at}sunstroke.sdsu.edu
Site-specific recombination (SSR) is involved in processes ranging from resolution of bacterial chromosome dimers to adeno-associated viral integration and is a versatile tool for mammalian genetics. The bacteriophage
J. Biol. Chem, 10.1074/jbc.M004679200
Submitted on May 30, 2000
Revised on August 2, 2000
Accepted on August 10, 2000
Characterization of a mutation of bacteriophage
integrase: putative role in core binding and strand exchange for a conserved residue
encoded site-specific recombinase Integrase (Int) is one of the best studied site-specific recombinases, and mediates recombination via 4 distinct pathways. We have characterized a mutant version of Int, IntT236I; this mutant can perform the bent-L pathway only, while the corresponding IntT236A mutant can perform both bent-L and excision pathways. Experiments involving both IntT236I and IntT236A show that the hydroxyl group of threonine is necessary for wild-type recombination. Substitution of the threonine by serine leads to nearly complete rescue of the mutant phenotypes. In addition, our data shows that the IntT236I mutant is defective partially due to obstructive steric interactions. Comparisons of crystal structures reveal that the threonine at residue 236 may play an important role in stabilizing recombination intermediates through solvent-mediated protein-DNA interactions at the core binding sites, and that the hydroxyl group is important for effective cleavage and Holliday junction formation. Our data also indicate that Int contacts the core sites differently in intermediates assembled in excisive versus bent-L recombination.
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