Oxidation of Neurospora crassa NADP-specific glutamate dehydrogenase by activated oxygen species.
The glutamine synthetase and the NADP-specific glutamate dehydrogenase activities of Neurospora crassa were lost in a culture without carbon source only when in the presence of air. Glutamine synthetase was previously reported to be liable to in vitro and in vivo inactivation by activated oxygen species. Here we report that NADP-specific glutamate dehydrogenase was remarkably stable in the presence of activated oxygen species but was rendered susceptible to oxidative inactivation when chelated iron was bound to the enzyme and either ascorbate or H2O2 reacted on the bound iron. This reaction gave rise to further modifications of the enzyme monomers by activated oxygen species, to partial dissociation of the oligomeric structure, and to precipitation and fragmentation of the enzyme. The in vitro oxidation reaction was affected by pH, temperature, and binding to the enzyme of NADPH. Heterogeneity in total charge was observed in the purified and immunoprecipitated enzymes, and the relative amounts of enzyme monomers with different isoelectric points changes with time of the oxidizing reaction.
Aguirre
J
J
Centro de Investigación sobre Fijación de Nitrógeno, Universidad Nacional Autónoma de México, Morelos.
Rodríguez
R
R
Hansberg
W
W
D000332:Aerobiosis; D000693:Anaerobiosis; D000818:Animals; D000906:Antibodies; D000936:Antigen-Antibody Complex; D001205:Ascorbic Acid / Q000494:pharmacology; D002417:Cattle; D003683:Dehydroascorbic Acid / Q000494:pharmacology; D004492:Edetic Acid / Q000494:pharmacology; D005969:Glutamate Dehydrogenase / Q000378:metabolism*; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007501:Iron / Q000494:pharmacology; D007700:Kinetics; D009249:NADP / Q000378:metabolism*; D009491:Neurospora; D009492:Neurospora crassa / Q000201:enzymology*; D010084:Oxidation-Reduction
D000906:Antibodies; D000936:Antigen-Antibody Complex; D009249:NADP; D004492:Edetic Acid; D006861:Hydrogen Peroxide; D007501:Iron; D005969:Glutamate Dehydrogenase; D001205:Ascorbic Acid; D003683:Dehydroascorbic Acid