Inhibition Kinetics of Cabbage Butterfly (Pieris rapae L.) Larvae Phenoloxidase Activity by 3-Hydroxy-4-Methoxybenzaldehyde Thiosemicarbazone
2007
Xue, C.B. | Luo, W.C. | Xie, X.Y. | Xiao, T. | Yan, L.
Phenoloxidase (PO) is a key enzyme in insect development, responsible for catalyzing the hydroxylation of tyrosine into o-diphenols and the oxidation of o-diphenols into o-quinones. In the present study, the kinetic assay in air-saturated solutions and the kinetic behavior of PO from Pieris rapae (Lepidoptera) larvae in the oxidation of l-tyrosine (a monophenol) and l-DOPA (l-3, 4-dihydroxyphenylalanine) (a diphenol) was studied. The inhibitory effects of 3-hydroxy-4-methoxybenzaldehyde thiosemicarbazone (3-H-4-MBT) on the monophenolase and diphenolase activities of PO were also studied. The results show that 3-H-4-MBT can inhibit both the monophenolase and diphenolase activities of PO. The lag period of l-tyrosine oxidation catalyzed by the enzyme was obviously lengthened and the steady-state activities of the enzyme sharply decreased. The inhibitor was found to be noncompetitively reversible with a K(I) (K(I) = K(IS)) of 0.30 μmol/L and an estimated IC50 of 0.14 +/- 0.02 μmol/L for monophenolase and 0.26 +/- 0.04 μmol/L for diphenolase. In the time course of the oxidation of l-DOPA catalyzed by the enzyme in the presence of different concentrations of 3-H-4-MBT, the rate decreased with increasing time until a straight line was approached. The microscopic rate constants for the reaction of 3-H-4-MBT with the enzyme were determined.
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