Mechanistic and stereochemical studies on the glycine reductase of Clostridium sticklandii.

Clostridial glycine reductase multienzyme complex which catalyses the reaction: Glycine + ADP + Pi + 2H leads to Acetate + ATP + NH3 was solubilised and fractionated essentially according to the method of Stadtman [T.C. Stadtman (1970) Methods Enzymol. 17A, 956--966] into two components: protein A and 'glycine reductase' fraction. A reconstituted system obtained by combining the two components in the presence of dithiothreitol catalysed the conversion of glycine into acetate concomitant with the phosphorylation of ADP to ATP. Using the reconstituted system, in which the unwanted enzyme activity catalyzing an exchange of the alpha hydrogen atoms of glycine with the protons of the medium had been greatly reduced, it was found that the conversion of (2RS)-[2-14C, 2-3H1]glycine (3H/14C = 7.16) into acetate (3H/14C = 7.03) was attended by the retention of both the C-2 hydrogen atoms of glycine. Conversion of (2S)-[2-2H1, 2-3H1]glycine and (2R)-[2-2H1, 2-3H1]glycine by the reconstituted system gave (2S)-acetate and (2R)-acetate respectively showing that the reductive deamination of glycine occurs through an inversion of configuration. The cumulative information available on the glycine reductase reaction is embodied in a hypothetical mechanism of action for the enzyme.