title: 9-azidoacridine, a new photoaffinity label for nucleotide- and aromatic-binding sites in proteins.
abstract: The effect of the photolytic reagent 9-azidoacridine, optionally 3H-labelled, was studied both kinetically and structurally on nine different enzymes, namely alpha-chymotrypsin, lactate dehydrogenase, glyceraldehyde 3-phosphate dehydrogenase, alcohol dehydrogenase, alanine dehydrogenase, D-amino acid oxidase, ribonuclease A, alkaline phosphatase and alpha-amylase. Dark inhibition was observed in several cases. The concentration of the inhibitor ranged from 0.2 microM to 0.67 microM and demonstrated competitive kinetics with nucleotide cofactors when present. All concentrations of inhibitor showed increased inhibition on photolysis. Examination of the oligopeptides from hydrolysis of the covalently 3H-labelled derivative in conjunction with known amino acid sequence and tertiary structure established that the primary site of interaction in those cases for which the tertiary structure was available involved the active-site region. The above results in conjunction with those obtained with the structural analogues 9-aminoacridine and 9-amino-1,2,3,4-tetrahydroacridine established that this reagent acts as a molecular probe of aromatic- and, in particular, nucleotide-binding sites. This reagent provides a further additional method for studying the nucleotide cofactor domain.
mesh_terms: D000345:Affinity Labels / Q000138:chemical synthesis / Q000494:pharmacology*; D000596:Amino Acids / Q000032:analysis; D001386:Azides / Q000138:chemical synthesis / Q000494:pharmacology*; D001665:Binding Sites; D004791:Enzyme Inhibitors; D007700:Kinetics; D009243:NAD / Q000378:metabolism*; D009249:NADP / Q000378:metabolism*; D009842:Oligopeptides / Q000032:analysis; D010446:Peptide Fragments / Q000032:analysis; D010782:Photolysis; D014357:Trypsin
chemical_list: D000345:Affinity Labels; D000596:Amino Acids; D001386:Azides; D004791:Enzyme Inhibitors; D009842:Oligopeptides; D010446:Peptide Fragments; D009243:NAD; C036356:9-azidoacridine; D009249:NADP; D014357:Trypsin