The role of electrostatic interactions in the mechanism of peptide bond hydrolysis by a Ser-Lys catalytic dyad

General-base catalysis in the active site of serine proteasesis carried out by the imidazole side chain of a histidine. Duringformation of the transition state, an adjacent carboxylic acidgroup stabilizes the positive charge that forms on the general-basecatalyst and as a result contributes several orders of magnitudeto the catalytic efficiency of these enzymes. In the recentlydiscovered family of self-cleaving proteins exemplified by theLexA repressor of Escherichia coli, instead of the imidazoleof a histidine, the active-site general-base catalyst was foundto be the -amino of a lysine. The considerably higher capacityof the lysine side chain for proton acceptance raises interestingquestions concerning the role of electrostatic interactionsin the mechanism of proton transfer by this highly basic group.The negative charge elimination studies described here and theireffects on the k_max and pK of LexA self-cleavage are consistentwith a model in which electrostatic interactions between anacidic side chain and the general-base catalyst form a barrierto proton transfer. The implications are that the -amino group,unlike the imidazole group, is capable of effecting proton transferwithout the intervention of a countercharge.