Y. Abdel-Aal, B. Hammock
1985
Citations
1
Influential Citations
63
Citations
Journal
Insect Biochemistry
Abstract
The nature of the inhibition of juvenile hormone esterase (JHE) from Trichoplusia ni (T. ni) by 3-octylthio-l,l,l-trifluoro-2-propanone (OTFP) was studied and the kinetic data presented are consistent with the mechanism of a slow and tight binding inhibitor. With the use of the above inhibitor, it was demonstrated that the classical methods, i.e. Lineweaver and Burk, which are based on the assumptions of a steady state are inadequate for determining the mechanism and the inhibition constants for the reaction of OTFP with JHE. The reaction was shown to he reversible based on the non-steady state kinetics with a dissocation constant (Ki) of ~ 1.2 x 10 -I° M. Values of 3.37 x 107 M -t min -t and 4.1 x 10 -3 min -t for the forward and reverse rate constants were calculated and indicate the nature of slow binding inhibition in comparison to ordinary substrate reactions. An application of Ackermann- Potter plots of tight binding inhibition was presented, and their usefulness in measuring the enzyme molar equivalency and its catalytic number towards JH-I and JH-III was evaluated. The molar equivalency of JHE in T. ni haemolymph which hydrolyzes JH-III (at a final concentration of 5.0 x 10 -6 M) at a rate of 33.8 nmol/min-ml was found to be 1.6 x 10 -6 M. The catalytic numbers for JH-I and JH-III were 37.1 and 19.4 min -~, respectively. These new kinetic parameters, in addition to the Michaelis constant (Km) and the maximum velocity (Vm~x) were applied to a study of enzyme--substrate specificity and an evaluation of the role of JHE in the regulation of JH-titre. Key Word Index: Trichoplusia ni, juvenile hormone esterase, inhibition, substrate specificity, kinetics, molarity, slow binding inhibitors, tight binding inhibitors, 3-octylthio-l,l,l-trifhioro-2-propanone