ELECTROSTATIC EVIDENCE FOR THE ACTIVATION OF THE GLUTATHIONE THIOL BYTYR7 IN PI-CLASS GLUTATHIONE TRANSFERASES

A number of spectrophotometric studies [Graminski, G. F., Kubo, Y. & A rmstrong, R. N. (1989) Biochemistry 28, 3562-3568; Liu, S., Zhang, P., Ji, X., Johnson, W. W., Gilliland, G. L. & Armstrong, R. N. (1992) J. BioL Chem. 267, 4296-4299] have recently shown that the glutathione ( GSH) thiol is deprotonated when it is in complex with glutathione S-tr ansferase. Different models have been proposed for the activation of t he glutathione S(gamma), all pointing out the key role of active-site residue Tyr7. It remains unclear, however, how Tyr7 is actually involv ed in this process. In this paper we present an analysis of the electr ostatic potential in the region of the active site of a pi-class GSH t ransferase. This analysis provides evidence that the titration behavio ur of the absorption band of the E . GSH complex with a pK between 6 a nd 7 [Liu, S., Zhang, P., Ji, X., Johnson, W. W., Gilliland, G. L. & A rmstrong, R. N. (1992) J. Biol. Chem. 267, 4296-4299] should rather be explained by the protonation/deprotonation equilibrium of Tyr7 than b y the protonation/deprotonation equilibrium of the GSH thiol group its elf. On the basis of this conclusion, a mechanism for activation of GS H is proposed: the Tyr7 OH group is deprotonated by the influence of t he protein charge constellation and the peptide dipoles. Thus it acts as a general base, promotes proton abstraction from the GSH thiol and creates a thiolate anion with high nucleophilic reactivity.