Kinetic evaluation of aminoethylisothiourea on mushroom tyrosinase activity.
This study demonstrates that aminoethylisothiourea (AET), a potent inhibitor of inducible nitric oxide synthase, is an irreversible competitive inhibitor of mushroom tyrosinase by chelation to the active site of tyrosinase when L-3,4-dihydroxyphenylalanine was assayed spectrophotometrically. The spectrophotometric recordings of the inhibition of tyrosinase by AET were characterized by the presence of a lag period prior to the attainment of an inhibited steady-state rate. The lag period corresponded to the time in which AET was reacting with the enzymatically generated o-quinone. Increasing AET concentrations provoked longer lag periods as well as a concomitant decrease in the tyrosinase activity. Both lag period and steady-state rate were dependent on AET, substrate, and tyrosinase concentrations. The inhibition of diphenolase activity of tyrosinase by AET showed positive kinetic cooperativity which arose from the protection of both substrate and o-quinone against inhibition by AET. The UV-visible spectrum of a mixture of tyrosinase and AET exhibited a characteristic shoulder peak ascribed to the chelation of AET to the active site of tyrosinase. Moreover, the presence of copper ions only partially prevented but not reverted mushroom tyrosinase inhibition when CuSO(4) was added to the assay medium on tyrosinase activity.
Li
Shu-Bai
SB
College of Chemistry, Chemical Engineering and Biotechnology, Donghua University, Shanghai 201620, China.
Nie
Hua-Li
HL
Zhang
Hai-Tao
HT
Xue
Yong
Y
Zhu
Li-Min
LM
D000363:Agaricales / Q000201:enzymology*; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D007700:Kinetics; D008956:Models, Chemical; D014442:Monophenol Monooxygenase / Q000378:metabolism*; D011809:Quinones / Q000378:metabolism; D000338:beta-Aminoethyl Isothiourea / Q000494:pharmacology*
D004791:Enzyme Inhibitors; D011809:Quinones; D000338:beta-Aminoethyl Isothiourea; D014442:Monophenol Monooxygenase