Purification and properties of two types of diphosphopyridine nucleotide-linked glycerol 3-phosphate dehydrogenases from chicken breast muscle and chicken liver.

We have purified and compared two different L-glycerol 3-phosphate:DPN-linked oxidoreductases (EC 1.1.1.8) from chicken, one from liver and the second from breast muscle. The crystalline enzyme from chicken breast muscle appears homogeneous by all tests applied. The purified glycerol j-phosphate dehydrogenase from chicken liver appears homogeneous by electrophoresis and sedimentation ultracentrifugation; however, it is not homogeneous as an antigen as determined by immunodiffusion. Both enzymes have molecular weights between 60,000 and 65,000 and appear to be composed of two identical subunits. The amino acid compositions are similar, with only glycine, glutamic acid plus glutamine, and arginine dsering by more than 3 residues per subunit. There are 2 tryptophan residues per subunit in each enzyme. These enzymes are markedly different by other criteria. Antisera, when added to each of these proteins, cross-reacts very weakly with the heterologous antigen. Electrophoresis at pH 8.6 shows that glycerol 3-phosphate dehydrogenase from chicken liver is negatively charged, whereas the corresponding breast muscle enzyme is positively charged. Rather striking differences are found in a comparison of the catalytic properties of glycerol 3-phosphate dehydrogenase from chicken liver and chicken breast muscle. The binding constants for oxidized and reduced diphosphopyridine nucleotide and the K, values for dihpdroxyacetone phosphate and glycerol 3-phosphate are, respectively, 5-, 25-, 5-, and lOO-fold lower for the liver enzyme than for its muscle homologue. l.f the catalytic properties in uifro are present under conditions in vivo, glycerol 3-phosphate dehydrogenase from chicken muscle would be essentially unable to catalyze the oxidation of glycerol j-phosphate. Peptide maps from tryptic digests of these two enzymes show little, if any, similarity, indicating that glycerol 3-phosphate dehydrogenases from chicken liver and chicken breast