Enzyme kinetics and substrate selectivities of rat glutathione S-transferase isoenzymes towards a series of new 2-substituted 1-chloro-4-nitrobenzenes
van der Aar, E.M.
te Koppele, J.M.
van Ommen, B.
van Bladeren, P.J.
Leiden/Amsterdam Ctr. for Drug Res., Department of Pharmacochemistry, Vrije Universiteit, De Boelelaan 1083, 1081 HV, Amsterdam, Netherlands TNO Preventie en Gezondheid TNO Voeding
1. Four different rat glutathione S-transferase (GST) isoenzymes, belonging to three different classes, were examined for their GSH conjugating capacity towards 11 2-substituted 1-chloro-4-nitrobenzene derivatives. Significant differences were found in their enzyme kinetic parameters K(m), k(cat) and k(cat)/K(m). 2. Substrates with bulky substituents on the ortho-position appeared to have high affinities (low K(m)'s) for the active site of the GST-isoenzymes, suggesting that there is sufficient space in this area of the active site. A remarkably high K(m) (low affinity) was found for 2-chloro-5-nitropyridine towards all GST-isoenzymes examined. 3. GST 3-3 catalysed the reaction between GSH and the substrates most efficiently (high k(cat)) compared with the other GST-isoenzymes. Moreover, GST 3-3 showed clear substrate selectivities towards the substrates with a trifluoromethyl-, chlorine- and bromine-substituent. 1-Chloro-2,4-dinitrobenzene and 2-chloro-5-nitrobenzonitrile were most efficiently conjugated by all four GST-isoenzymes examined. 4. When the rate of the conjugation reactions was followed, a linear increase of formation of GS-conjugate could be seen for 2-chloro-5-nitrobenzonitrile during a much longer period of time than for 1-chloro-2,4-dinitrobenzene with all GST-isoenzymes examined. Therefore, it is suggested that 2-chloro-5-nitrobenzonitrile might be recommended as an alternative model substrate in GST-research. Chemicals/CAS: Dinitrochlorobenzene, 97-00-7; Glutathione Transferase, EC 188.8.131.52; Isoenzymes
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Xenobiotica, 26 (2), 143-155