The Nucleotide and Acyl Group Content of Native Rabbit Muscle Glyceraldehyde 3-Phosphate Dehydrogenase

Glyceraldehyde 3-phosphate dehydrogenase (GP-dehydrogenase) isolated from rabbit muscle by the procedure of Ferdinand (Biochem. J., 92, 578 (1964)), has two previously unreported properties. 1. The coenzyme-binding sites usually filled with NAD+ in native GP-dehydrogenase contain instead the NAD+ degradation product, ADP-ribose. 2. Several of the four active sites contain covalently bound 3-phosphoglyceroyl group. The enzyme isolated by the Ferdinand procedure is in reality the acyl enzyme "intermediate" in the physiological oxidative phosphorylation of glyceraldehyde 3-phosphate. The occurrence of ADP-ribose appears to be a consequence of the isolation procedure in vitro, but the recovery of acyl enzyme suggests that GP-dehydrogenase is in an acylated state in vivo. The data required to estimate the sarcoplasmic concentrations of NAD+, NADH, the glycolytic intermediates, and the glycolytic enzymes are available in the literature. These estimates and the known effect of acylation on the GP-dehydrogenase-NAD+ affinity suggest two physiological regulatory roles for acyl-GP-dehydrogenase in muscle. One function is the regulation of coenzyme activities and oxidation-reduction potential as the affinity of enzyme for NAD+ changes with degree of acylation, itself dependent on the metabolic state of the tissue. The other is the "buffering" by acyl enzyme of the more dilute glycolytic intermediates during rapid changes in metabolic state. Although these roles must be considered hypothetical, the finding in skeletal muscle of levels of acylated GP-dehydrogenase much higher than those of many glycolytic intermediates should lead to revision of the accepted glycolytic pathway to include 3-phosphoglyceroyl-GP-dehydrogenase as a significant metabolite.