HPLC study of tyrosinase inhibition by thiopronine.
Thiopronine (N-2-mercaptopropionyl-glycine, NMPG) inhibits the o-dihydroxy-phenolase activities of mushroom tyrosinase. When d,l-3-4-dihydroxyphenylalanine (DOPA) is employed as substrate, the inhibition was found to be a competitive-type with K(i) of 0.95 micro m. We found in addition that thiopronine interacts with the enzymatic generated product (o-quinone) to form a colourless conjugate compound causing an apparent inhibition. These data suggest that thiopronine inhibits mushroom tyrosinase activity in two ways: (1) by forming an adduct with dopaquinone; and (2) by direct interaction with the enzyme probably towards the copper (II) present in the active site or cysteine-rich domains. This finding was indicated by the presence of a lag period prior to the attainment of an inhibited steady-state rate. Both lag period and steady-state rate were dependent on thiopronine and substrate concentrations. An increase of thiopronine concentration causes longer lag periods as well as a concomitant decrease in the tyrosinase activity. The presence of an excess of copper (II) reverses the inhibition exerted by thiopronine.
Girelli
A M
AM
Dipartimento di Chimica, Università 'La Sapienza', P.le A. Moro 5, 00185 Roma, Italy. girelli@uniroma1.it
Mattei
E
E
Messina
A
A
D000363:Agaricales / Q000201:enzymology; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D014442:Monophenol Monooxygenase / Q000037:antagonists & inhibitors*; D013056:Spectrophotometry, Ultraviolet; D008625:Tiopronin / Q000494:pharmacology*
D004791:Enzyme Inhibitors; D008625:Tiopronin; D014442:Monophenol Monooxygenase