In vitro analysis of mutant LexA proteins with an increased rate of specific cleavage

Analysis of Escherichia coli RecA Interactions with LexA, λ CI, and UmuD by Site-Directed Mutagenesis of recA

“…Our evidence with LexA favors a conformational model in which RecA stabilizes a reactive conformation of LexA (44). However, it remains possible that RecA makes a more direct contribution to the chemistry of bond breakage.…”

A monocysteine approach for probing the structure and interactions of the UmuD protein

“…In their studies of the cleavage of LexA repressor, Roland et al (49) reported mutations in LexA which resulted in hypercleavable repressors, presumably by causing a conformation that is competent for cleavage. They proposed that RecA favors this conformation and thus increases the rate of reaction.…”

“…For the related proteins LexA and X repressor, the third possibility is particularly likely. Roland et al (49) and Slilaty and Little (54) hypothesized that the region containing the nucleophilic serine in LexA, Ser-119, is probably not well exposed to solvent. This conclusion is based on the finding that previous attempts to inhibit LexA autodigestion with the serine (54) and that a much higher concentration of diisopropyl fluorophosphate (20 mM compared with 1 mM previously tried) is required to modify 50% of the LexA in the 10-min incubation period and result in an inhibitory effect on autodigestion (48).…”

LexA cleavage and other self-processing reactions

“…We reasoned (34,41) that analysis of mutant LexA proteins with an increased rate of cleavage might help identify the rate-limiting step or steps in cleavage. The rate of any chemical reaction is determined by the free energy difference between the ground state and the transition state.…”

“…This model was suggested by the properties of a new class of1exA mutations, termed hypercleavable or Inds mutations, that confer increases in the rate of specific cleavage (34,41 (18). A truncated X CI substrate is not cleaved by either enzyme.…”