INHIBITION OF CHICKEN LIVER CARBOXYLESTERASE BY ACTIVATED CARBONYLS AND CARBONYL HYDRATES

Identical k(cat) values (approximate to 40 s(-1)) are obtained for the chicken liver carboxylesterase catalyzed hydrolysis of phenyl, p-nitr ophenyl and o-nitrophenyl benzoates providing support for the involvem ent of an acyl-enzyme pathway, with the rate-limiting deacylation of a common benzoyl-enzyme intermediate. Chicken liver carboxylesterase ca talyzed fragmentation of (E)-benzilmonoxime O-2,4-dinitrophenyl ether shows a pH dependence on a group active in the free base form with a p K(a)' approximate to 5.0. The K-i-pH profile for benzil inhibition sho ws a dependence on a similar group with a pK(a)' = 5.4. The reactions between chicken liver carboxylesterase and the hydrated aldehyde, chlo ral hydrate, have shown this compound to be at once a substrate and po tent inhibitor of the enzyme. The kinetics of inhibition are consisten t with a mechanism in which the bound hydrate is first dehydrated in a rate-limiting step catalyzed by the enzyme. Nucleophilic attack by th e active-site serine on the parent aldehyde produces a hemiacetal addu ct. The K-i value for chloral hydrate inhibition calculated from the k inetic analysis (90 nM) compares favourably with the value measured fr om the steady-state kinetics (87 nM).