Disorder-to-order transition of the active site of human class Pi glutathione transferase, GST P1-1

Glutathione transferases comprise a large family of cellular detoxification enzymes that function by catalyzing the conjugation of glutathione (GSH) t o electron-deficient centers on carcinogens and other toxins. NMR methods h ave been used to characterize the structure and dynamics of a human class p i enzyme, GST P1-1, in solution. Resonance assignments have been obtained f or the unliganded enzyme and the GSH and S-hexylglutathione (GS-hexyl) comp lexes. Differences in chemical shifts between the GSH and GS-hexyl complexe s suggest more extensive structural differences between these two enzyme-li gand complexes than detected by previous crystallographic methods. The NMR studies reported here clearly show that an alpha -helix (alpha2) within the GSH binding site exists in multiple conformations at physiological tempera tures in the absence of ligand. A single conformation of alpha2 is induced by tile presence of either GSH or GS-hexyl or a reduction in temperature to below 290 K. The large enthalpy of the transition (similar to 150 kJ/mol) suggests a considerable structural rearrangement of the protein. The Gibbs free energy for the transition to the unfolded form is on the order of -4 t o -6 kJ/mol at physiological temperatures (37 degreesC). This order-to-diso rder transition contributes substantially to the overall thermodynamics of ligand binding and should be considered in the design of selective inhibito rs of class pi glutathione transferases.