Purification and some properties of mitochondrial glutamate:glyoxylate aminotransferase and mechanism of its involvement in glycolate pathway in Euglena gracilis z.

Euglena contains glutamate:glyoxylate aminotransferase (GGT) both in mitochondria and in cytosol. Both isoforms were separated from each other by DEAE-cellulose chromatography. The mitochondrial enzyme had an apparent Km of 1.9 mM for glutamate and the cytosolic enzyme 52.6 mM. Mitochondrial GGT was further purified by ammonium sulfate fractionation, isoelectric focusing, and gel chromatography. It had a molecular weight of 141,000 and an isoelectric point of pH 4.88; the optimum pH was 8.5. Its apparent Km values for glutamate and for glyoxylate were 2.0 and 0.25 mM, respectively. In addition to glutamate, mitochondrial GGT used 5-hydroxytryptophan, tryptophan, and cysteine as amino donors in the transamination to glyoxylate. Alanine did not support the activity. The relative activity of the enzyme for amino acceptors on the transamination from glutamate was 4-hydroxyphenylpyruvate greater than phenylpyruvate greater than glyoxylate greater than hydroxypyruvate. Pyruvate and 2-oxoglutarate were not used in the reaction. Evidence that GGT functions mainly in the irreversible transamination between glutamate and glyoxylate is presented. The functional significance of GGT in the glycolate pathway of Euglena is also discussed.