Mutation of an evolutionarily conserved tyrosine residue in the active site of a human class Alpha glutathione transferase

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Abstract
Human class Alpha glutathione transferase (GST) A1—1 has been subjected to site‐directed mutagenesis of a Tyr residue conserved in all classes of cytosolic GSTs. The change of Tyr8→Phe lowers the specific activities with three substrates to 2–8% of the values for the wild‐type enzyme. The changes in the kinetic parameters k cat/K max, V max and S0.5 show that the decreased activities are partly due to a reduced affinity for glutathione. The effect is reflected in lowered k cat values, suggesting that the hydroxyl group of Tyr8 is involved in the activation of glutathione. The proposal of such a role for the Tyr residue has support from the 3D structure of a pig lung class Pi GST [Reinemer et al. (1991) EMBO J. 10, 1997–2005]. Thus, Tyr8 appears to be the first active site residue established as participating in the chemical mechanism of a GST.