Serine transhydroxymethylase: Mechanism of aldolase activation by folate

Abstract

Affinity chromatographic methods have been developed to purify beef liver serine transhydroxymethylase. This enzyme catalyzes both the transfer of aldehydic groups from β-hydroxy amino acids to tetrahydrofolate and the cleavage of β-hydroxy amino acids yielding the free aldehyde. Tetrahydrofolate, folate, and a quinazoline analog of isofolate were found to be activators of the β-phenylserine aldolase reaction catalyzed by serine transhydroxymethylase. Activation by folate was maximal at 20 μ m and higher concentrations diminished the activation. Evidence is presented suggesting that folate does not activate by providing an acceptor site for the aldehydic groups. Equilibrium binding studies showed that folate and tetrahydrofolate can bind the enzyme with essentially the same affinity. Double-reciprocal plots with β-phenylserine from steady-state kinetic experiments did not yield a 1 v 1 , intercept effect except at high folate concentrations. A mechanism is proposed in which folate binds readily to the enzymic active site, facilitating β-phenylserine binding. Folate is subsequently lost, at least partially, prior to product release and complete enzymic turnover.