The essential histidine residues of bovine plasma amine oxidase.
Ethoxyformic acid anhydride and photooxidation have been used to study the function of histidine residues in bovine plasma amine oxidase. Ethoxyformic acid anhydride at pH 6.1 reacted with nearly all of the histidine residues in the enzyme in 15 min but complete enzyme inactivation occurred in several minutes. The concentration of the reagent which caused 50% inhibition was 2.2-10(-5) M under the conditions of the experiment. The diamine oxidases, Aspergillus niger and pea seedling amine oxidases were also inhibited by ethoxyformic acid anhydride. The concentrations of reagent required for 50% inhibition were 6.6-10(-5) and 3.3-10(-4) M, respectively, for the two enzymes. NH2OH could not be used to regenerate the reacted histidine residues since NH2OH itself inhibited the enzyme. Photooxidation in the presence of 0.001% Rose Bengal at pH 7.0 also inactivated bovine plasma amine oxidase. Histidine was the only amino acid destroyed by photooxidation. About six histidine residues were destroyed but in the presence of the substrate kynuramine, two less histidine residues were destroyed. Since lysine which is neither a substrate nor inhibitor of the enzyme did not protect the enzyme from photooxidation, it was concluded that two histidine residues, one in each sub-unit of the enzyme are essential for activity.
D000596:Amino Acids / Q000032:analysis; D000812:Anhydrides / Q000494:pharmacology; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D005561:Formates / Q000494:pharmacology; D006639:Histidine / Q000032:analysis*; D007700:Kinetics; D010084:Oxidation-Reduction; D000587:Oxidoreductases Acting on CH-NH Group Donors / Q000032:analysis / Q000097:blood*; D010777:Photochemistry; D011485:Protein Binding; D012395:Rose Bengal / Q000494:pharmacology
D000596:Amino Acids; D000812:Anhydrides; D005561:Formates; D012395:Rose Bengal; D006639:Histidine; D000587:Oxidoreductases Acting on CH-NH Group Donors