Kinetics of rat brain soluble catechol-O-methyltransferase and its inhibition by substrate analogues.
1. The initial rates and inhibition of rat brain catechol-O-methyltransferase were studied. Double reciprocal plots of initial rates versus either S-adenosyl-L-methionine or 3,4-dihydroxybenzoic acid, in the absence of product, gave a series of lines intersecting to the left of the ordinate. 2. Inhibition in the presence of S-adenosyl-L-homocysteine was competitive but in the presence of vanillic acid was non-competitive if S-adenosyl-L-methionine was the varied substrate. 3. When 3,4-dihydroxybenzoic acid was the varied substrate, both S-adenosyl-L-homocysteine and vanillic acid gave rise to a non-competitive inhibition. 4. The initial rate and product inhibition patterns were consistent with an ordered BiBi mechanism with S-adenosyl-L-methionine being the first substrate and 3,4-dihydroxybenzoic acid the second substrate to combine with the enzyme. 5. In addition, these results suggest that vanillic acid is the first product and S-adenosyl-L-homocysteine the second product to dissociate from the enzyme. 6. The substrate analogues salsolinol and 3-carboxysalsolinol were competitive inhibitors with respect to 3,4-dihydroxybenzoic acid but were non-competitive with respect to S-adenosyl-L-methionine. For enzymes with an ordered mechanism an uncompetitive inhibition would be expected. 7. A possible explanation is that both substrate analogues can combine with either free enzyme with lower affinity or with an intermediary enzyme form with much greater affinity. 8. A scheme which is consistent with the data is presented.
D000818:Animals; D001921:Brain / Q000201:enzymology*; D065098:Catechol O-Methyltransferase Inhibitors; D055598:Chemical Phenomena; D002621:Chemistry; D066298:In Vitro Techniques; D007546:Isoquinolines / Q000494:pharmacology*; D007700:Kinetics; D008297:Male; D051381:Rats; D011919:Rats, Inbred Strains; D012490:Salsoline Alkaloids / Q000494:pharmacology*; D012995:Solubility; D013379:Substrate Specificity
D065098:Catechol O-Methyltransferase Inhibitors; D007546:Isoquinolines; D012490:Salsoline Alkaloids; C011704:carboxysalsolinol; C036617:salsolinol