[Structural role of histidine residues in NAD(P)-glutamate dehydrogenase from the bovine liver].
Photooxidation of bovine liver glutamate dehydrogenase (GDH, EC 1.4.1.3) in the presence of methylene blue at a low light intensity occurs in two stages. At the first stage, the duration of which depends on temperature and dye concentration, a slight activation is observed simultaneously with the oxidation of two histidine residues. At the second stage, the inactivation is concomitant with the oxidation of three histidine and one tryptophan residues. The inactivation is a first order reaction (k = 3,22 X 10(-2) min-1) and is correlated with changes in the circular dichroism spectra. These data testify to the structural role of histidine residues in the GDH molecule. The kinetic behaviour of GDH during its modification with diethylpyrocarbonate (DEP) depends on pH and the reagent concentration. Four histidine residues undergo carbethoxylation at pH 6.0 and 7.5, but the modification rate is much higher at pH 7.5. At low DEP concentrations, a remarkable activation is observed with a simultaneous modification of one histidine residue, which is independent of pH. At high DEP concentrations, a rapid inactivation takes place at pH 7.5. Treatment of the carbethoxylated inactive enzyme with hydroxylamine results in the deacylation of histidine residues without any noticeable reactivation. The data on the combined effect of DEP and pyridoxal-5'-phosphate suggest that GDH inactivation by DEP at pH 7.5 is a result of modification of an essential epsilon-NH2 group of lysine-126.
D000818:Animals; D002417:Cattle; D002942:Circular Dichroism; D004047:Diethyl Pyrocarbonate / Q000494:pharmacology; D005969:Glutamate Dehydrogenase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006639:Histidine / Q000032:analysis; D066298:In Vitro Techniques; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D009249:NADP / Q000378:metabolism*; D010084:Oxidation-Reduction; D010777:Photochemistry
D006639:Histidine; D009249:NADP; D005969:Glutamate Dehydrogenase; D004047:Diethyl Pyrocarbonate
Strukturnaia rol' ostatkov gistidina NAD(P)-glutamatdegidrogenazy pecheni byka.