title: Mutant LexA proteins with an increased rate of in vivo cleavage.
abstract: LexA repressor of Escherichia coli is inactivated by a specific cleavage reaction that requires activated RecA protein in vivo. This cleavage reaction can proceed in vitro in the presence of activated RecA or as an intramolecular RecA-independent reaction, termed autodigestion, that is stimulated by alkaline pH. Here we describe a set of LexA mutant proteins that undergo a greatly increased rate of specific cleavage in vivo, compared with wild-type LexA. Efficient in vivo cleavage of these mutant proteins also took place without RecA. Several lines of evidence suggest that cleavage occurred via a mechanism similar to autodigestion. These mutations changed Gln-92, which lies near the cleavage site, to tyrosine, phenylalanine, or tryptophan. The latter mutation increased the rate of cleavage approximately 500-fold. These findings imply that the rate of wild-type LexA cleavage has been optimized during evolution to make the SOS system properly responsive to DNA-damaging treatments. Availability of these mutants will aid in the understanding of rate-limiting steps in intramolecular reactions.
authors.affiliation: Department of Biochemistry, University of Arizona, Tucson 85721.
authors.forename: M H
authors.initials: MH
authors.lastname: Smith
authors.forename: M M
authors.initials: MM
authors.lastname: Cavenagh
authors.forename: J W
authors.initials: JW
authors.lastname: Little
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005838:Genotype; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D010957:Plasmids; D012097:Repressor Proteins / Q000378:metabolism*; D012697:Serine Endopeptidases; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism; D000099160:LexA Repressor Protein
chemical_list: D001426:Bacterial Proteins; D012097:Repressor Proteins; D012697:Serine Endopeptidases; D001616:beta-Galactosidase; D000099160:LexA Repressor Protein