Affinity labeling of histidine and lysine residue in the adenosine deaminase substrate binding site.
1. Adenosine deaminase was inactivated by 9-(4-bromoacetamidobenzyl)-adenine (I) and 9-(2-bromoacetamidobenzyl)adenine (II), two affinity labels. 2. The stoichiometry of the reaction with reagent II is reported: 1 mol reagent is bound per mol inactive enzyme. Amino acid analysis of the 6 N HCl hydrolyzate of the inactive enzyme identified CM-histidine as the main alkylation product. This is the first evidence of the presence of a histidine in the active site region. 3. The alkylation rate and involved amino acid residues were studied for both reagents I and II, at pH 8 and 5.5. The particular reactivity of a lysine near or in the active site is discussed.
D000225:Adenine / Q000031:analogs & derivatives*; D058892:Adenosine Deaminase Inhibitors; D000345:Affinity Labels; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D002850:Chromatography, Gel; D006639:Histidine; D006863:Hydrogen-Ion Concentration; D008239:Lysine; D009700:Nucleoside Deaminases / Q000037:antagonists & inhibitors*
D058892:Adenosine Deaminase Inhibitors; D000345:Affinity Labels; C020984:9-(2-bromoacetamidobenzyl)adenine; D006639:Histidine; D009700:Nucleoside Deaminases; D000225:Adenine; D008239:Lysine