Inhibition of 2-amino-3-methylimidazo[4,5-f]quinoline (IQ)-DNA binding by chlorophyllin: studies of enzyme inhibition and molecular complex formation.
Chlorophyllin (CHL), a copper/sodium salt of chlorophyll used in the treatment of geriatric patients, is an anti-mutagen that has been demonstrated to inhibit carcinogen--DNA binding in vivo. To study the mechanism of inhibition, the microsomal metabolism of 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and the kinetics of IQ--DNA binding were investigated in the presence and absence of CHL. In time-course studies, CHL produced greater than 80% inhibition of IQ--DNA binding and blocked the metabolism of IQ, such that 80% of the initial dose of carcinogen was recovered unmetabolized from the incubations after 1 h. Kinetic constants were determined for the in vitro DNA binding reaction, with the reaction rate measured as 'pmol IQ bound/mg DNA/min/mg microsomal protein'. Without altering V(max), the Km of the IQ--DNA binding reaction was increased by CHL, and the replot of Km/V(max) versus CHL concentration yielded a straight line with an inhibitor constant of 58.3 microM CHL. Spectrophotometric studies provided evidence in vitro for the formation of a non-covalent complex between CHL and IQ. The CHL--IQ complex had a stoichiometric ratio of 2:1 (mole ratio method) and an apparent dissociation constant from the Benesi-Hilderbrand plot of 1.41 x 10(-4)M at pH 7.4. These results are discussed in the context of a CHL inhibitory mechanism involving enzyme inhibition and molecular complex formation.
Dashwood
R
R
Department of Environmental Biochemistry, University of Hawaii, Honolulu 96822.
Guo
D
D
D000818:Animals; D000891:Anti-Infective Agents, Local / Q000494:pharmacology; D001571:Benzoflavones / Q000494:pharmacology; D002273:Carcinogens / Q000378:metabolism*; D002735:Chlorophyllides / Q000494:pharmacology*; D004247:DNA / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology*; D011804:Quinolines / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D013053:Spectrophotometry; D019324:beta-Naphthoflavone
D000891:Anti-Infective Agents, Local; D001571:Benzoflavones; D002273:Carcinogens; D002735:Chlorophyllides; D011804:Quinolines; C029216:2-amino-3-methylimidazo(4,5-f)quinoline; D019324:beta-Naphthoflavone; D004247:DNA; C007020:chlorophyllin