Brain histamine N-methyltransferase purification, mechanism of action, and inhibition by drugs

Abstract

Histamine N-methyltransferase (E.C., 2.1.1.8; HMT) has been purified approximately 200-fold with about 20 per cent yield from guinea pig brain. The purification steps involved centrifugation at 225,000 g for 12hr, calcium phosphate gel adsorption, DEAE-cellulose chromatography and final chromatography on hydroxylapatite. The enzyme obtained is the most highly purified brain HMT so far prepared. It is specific for histamine with a K m of (4.30 ± 0.14) × 10 −5 M; it fails to methylate norepinephrine, serotonin and betazole. a histamine analogue. The kinetics and mechanism of action of histamine N-methyltransferase were investigated. Initial velocity studies at low substrate concentrations suggested that this methyltransferase action proceeded through two half-reactions in a ping pong mechanism with the intermediate formation of a methylated enzyme. This type of mechanism was also supported by results of product inhibition studies, in which methylhistamine was found to inhibit the enzyme competitively with respect to S-adenosylmethionine and noncompetitively with respect to histamine. Several drugs and chemicals were found to inhibit the enzyme; they included mercurial diuretics, antimalarials, antihistaminics, local anesthetics and ethylamine derivatives. The inhibition by each of these compounds, usually at a concentration of 10 −4 M or lower, was competitive with respect to histamine and appeared to be mixed with respect to S-adenosylmethionine. These patterns of inhibition also have a strong support for the ping pong mechanism.