Effect of captopril on mushroom tyrosinase activity in vitro

The study presented here demonstrates that the antihypertensive drug captop ril ([2S]-N-[3-mercapto-2-methylpropionyl]-L-proline) is an irreversible no n-competitive inhibitor and an irreversible competitive inhibitor of the mo nophenolase and diphenolase activities of mushroom tyrosinase when L-tyrosi ne and L-DOPA were assayed spectrophotometrically in vitro, respectively. C aytopril was rendered unstable by tyrosinase catalysis because of the inter action between the enzymatic-generated product (omicron -quinone) and capto pril to give rise to a colourless conjugate. Therefore, captopril was able to prevent melanin formation. The spectrophotometric recordings of the inhi bition of tyrosinase by captopril were characterised by the presence of a l ag period prior to the attainment of an inhibited steady state rate. The la g period corresponded to the time in which captopril was reacting with the enzymatically generated o-quinone. Increasing captopril concentrations prov oked longer lag periods as well as a concomitant decrease in the tyrosinase activity. Both lag period and steady state rate were dependent of captopri l, substrate and tyrosinase concentrations. The inhibition of both monophen olase and diphenolase activities of tyrosinase by captopril showed positive kinetic co-operativity which arose from the protection of both substrate a nd o-quinone against inhibition by captopril. Inhibition experiments carrie d out using a latent mushroom tyrosinase demonstrated that captopril only b ound the enzyme at its active site. The presence of copper ions only partia lly prevented but not reverted mushroom tyrosinase inhibition. This could b e due to the formation of both copper-captopril complex and disulphide inte rchange reactions between captopril and cysteine rich domains at the active site of the enzyme. (C) 2001 Elsevier Science B.V. All rights reserved.