Vitamin B-6-catalyzed beta-elimination of serine and O-phosphoserine. Qualitative and quantitative aspects of catalytic influences at the rate-limiting step, a comparison with the rate of enzymatic beta-elimination.

Abstract

The overall reaction rates for the beta-elimination of serine and O-phosphoserine, catalyzed by various vitamin B-6 analogs (pyridoxal 5'-phosphate, 5'-deoxypyridoxal and N-methylpyridoxal 5'-phosphate) in the presence or absence of Cu2+ ions, are determined. The comparison of the pH-dependence of the molar activities of the three vitamin B-6 aldehydes in beta-elimination of serine enables the characterization of the different active Schiff base species and the single catalytic events. The Schiff base which has a positive charge on the pyridine ring nitrogen and a fully ionized phosphate group shows the highest molar activity. The phosphate group acts as an intramolecular general base catalyst, most probably at the alpha-carbon proton of the amino acid. Furthermore general acid catalysis by buffer species occurs at the beta-hydroxy group serine. These facts together provide a kinetically unambiguous description of the mechanism of the reaction: the removal of the proton at the alpha-carbon atom of serine is the rate-limiting step and is followed by the more rapid elimination of the b-hydroxy group of serine. The forward rate constant of the rate-limiting step is calculated for each of the reactions mentioned. The rate constants are compared with respect to the effectiveness of the individual catalytic components in the vitamin B-6-dependent beta-elimination. For optimal conditions the reaction of O-phosphoserine is faster by a factor of 10(4) in the velocity of the beta-elimination than the corresponding acid-catalyzed beta-elimination of serine. For the eliminations at the alpha- and beta-carbon atoms of O-phosphoserine in vitamin B-6-catalysed reactions a common transition state is discussed. From a comparison of the fastest vitamin beta-6-dependent model reaction with the rate of an enzymatic beta-elimination it is suggested that for those beta-elininating enzymes where the rate-limiting step is the same as in the model, the catalytic components mentioned could suffice to explain the velocity of the rate-limiting step.