Allosteric Effects in Nicotinamide Adenine Dinucleotide Phosphate-specific Glutamate Dehydrogenase from Neurospora

Abstract

Abstract Neurospora nicotinamide adenine dinucleotide phosphate-specific glutamate dehydrogenase is predominantly in an inactive form at pH 7.2, is fully active above pH 8.0, and is partially active at intermediate pH values. Between pH 7.0 and 7.4, and in the absence of NADPH, the enzyme is activated by the substrates 2-oxoglutarate and glutamate, as well as by the nonsubstrates citrate, isocitrate, succinate, malate, fumarate, oxalacetate, or ethylenediaminetetraacetate. Curves relating activation to activator or substrate concentration were found to be sigmoid, indicating cooperation between activating molecules. At lower pH values the addition of NADPH results in nearly complete inactivation. Under these conditions activation requires 2-oxoglutarate specifically; glutamate or nonsubstrate acids have no activating effect. However, 2-oxoglutarate activates even better than in the absence of NADPH. NADP+ causes some activation by itself and enhances activation by 2-oxoglutarate, glutamate, and succinate. It is concluded that the enzyme has allosteric properties, the possible regulatory significance of which is briefly discussed. Properties previously reported for certain mutant varieties of this enzyme obtained from amination-deficient (am) mutants and their partial revertants are now seen as distorted manifestations of the normal allosteric interactions reported here for the enzyme obtained from a wild type strain.