Abstract
The reactivity of a number of catechols and catecholamines with regard to the enzymatic O-methylation by catechol-O-methyltransferase (COMT) was studied. The reaction was carried out in a vial at a temperature of 37 degrees C, the vial contained a certain concentration of one catechol(amine), catechol-O-methyltransferase (the enzyme), S-adenosylmethionine (the methyldonor), MgCl2 (a cofactor) and buffer pH = 7.85. After certain time intervals samples were taken, the reaction was stopped in acid. The catechol(amine) concentration decrease and the product concentration increase were determined by injecting the samples directly into an HPLC connected with a fluorimeter, giving the opportunity of estimating the mass balance. Vmax, Km, C3/C4 ratio (= ratio of 3-O- and 4-O-methylated product formed) and the reaction rate at low substrate (catecholamine) concentration (= Vmax. [S]/Km)-which appears to be related to log P - are given. It is conjectured that V at low substrate concentration is especially determined by the polarity of the catechol(amine) while Vmax is primarily determined by other physico-chemical properties like steric conformation (L-dopa vs DL-dopa; L-adrenaline vs DL-adrenaline).
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