Inhibition of human placenta glutathione transferase P1-1 by the antibiotic calvatic acid and its diazocyanide analogue--evidence for multiple catalytic intermediates.
ABSTRACT The inhibition mechanism of the dimeric human placenta glutathione transferase (GST) P1-1 by calvatic acid and the reaction intermediates, i.e. the diazocyanide analogue of calvatic acid, has been investigated at pH 7.0 and 30.0 degrees C. Experiments performed at different molar ratios of inhibitor/GST P1-1 indicate that 1 mol calvatic acid inactivates 1 mol GST P1-1, containing two catalytically equivalent active sites. However, 2 mol of the diazocyanide analogue of calvatic acid inactivate 1 mol GST P1-1. Two disulfide bridges/dimer, probably between Cys47 and Cys101, have been formed during the reaction of GST P1-1 with calvatic acid and its diazocyanide analogue. The apparent second-order rate constants for GST P1-1 inactivation by calvatic acid and its diazocyanide analogue are 2.4+/-0.3 M(-1) s(-1) and (8.5+/-0.7) x 10(3) M(-1) s(-1), respectively. The reaction of calvatic acid with free L-cysteine can be described by a simple process with an apparent second-order rate constant of (5.0+/-0.4) x 10(1) M(-1) s(-1). In contrast, a transient species occurs during the reaction of the diazocyanide analogue of calvatic acid with free L-cysteine. Kinetics may be described by a second-order process [the rate constant being (8.0+/-0.5) x 10(3) M(-1) s(-1)] followed by a first-order decay [the rate constant corresponding to (1.2+/-0.1) x 10(1) s(-1)]. Calvatic acid represents an enzyme inhibitor acting much slower than its reaction intermediates (i.e. its diazocyanide analogue).
- European Journal of Cancer 07/1996; 32A(6):967-78. · 5.06 Impact Factor
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ABSTRACT: The effect of phenylazoxycyanide and calvatic acid, its reference antibiotic, on some functions of tubulin obtained from different sources has been studied. Our purpose was to establish a possible correlation between the antitumour activity of these drugs and their antimicrotubular action. Microtubules are subcellular structures involved in proliferation and maintenance of the cell shape and probably in malignant transformation; indeed most antimitotic drugs influence the stability of microtubules through the interaction with tubulin, their main protein. In this work we found phenylazoxycyanide impairs, more than calvatic acid, polymerization of purified tubulin from calf brain. It also damages, in a dose-dependent manner, colchicine-binding ability of tubulin derived from rat liver and AH-130 Yoshida ascite hepatoma cells. Compounds displaying an azoxycyano group may represent new antimicrotubular agents and their effect could be modulated by the different polarity and structural characteristic of the molecule.Biochemical Pharmacology 05/1989; 38(7):1121-4. · 4.58 Impact Factor
- Journal of Biological Chemistry 12/1974; 249(22):7130-9. · 4.65 Impact Factor