Substrate proton exchange catalyzed by gamma-cystathionase.

Pulsed Fourier transform proton magnetic resonance was used to study the labilization of protons of various L-amino acids by the enzyme gamma-cystathionase. In the course of the normal reaction, the enzyme labilizes the alpha and beta protons of the substrate, L-homoserine, and promotes elimination of the gamma substituent. It was found that gamma-cystathionase also catalyzes the exchange of the alpha and beta protons of L-amino acids which cannot undergo elimination reactions, but are competitive inhibitors of the enzyme. Both beta protons of L-alpha-aminobutyrate, although not stereochemically equivalent, were exchanged at equal rates, whereas selectivity was shown for one of the beta hydrogens when the carbon length was increased. The data also show that beta-proton exchange cannot occur without alpha-proton exchange. The rate of alpha-proton exchange from amino acids containing a terminal hydroxyl group at the beta, gamma, or lambda carbon is greater than from the corresponding unsubstituted amino acid. Exchange rates of the alpha proton for the inhibitors examined vary from one-seventh that of the normal enzymatic reaction to approximately the same rate as that for the elimination reaction with homoserine. An active site with two areas of substrate-enzyme interaction is proposed. One site contains pyridoxal 5'-phosphate and the base or bases involved in alpha- and beta-proton exchange; the second site contains a base which normally facilitates removal of the gamma substituent and can interact with the gamma and lambda carbons of the substrate molecule.