Glutathione transferases comprise a large family of cellular detoxification enzymes that function by catalyzing the conjugation of glutathione (GSH) to electron-deficient centers on carcinogens and other toxins. NMR methods have been used to characterize the structure and dynamics of a human class pi enzyme, GST P1-1, in solution. Resonance assignments have been obtained for the unliganded enzyme and the GSH and S-hexylglutathione (GS-hexyl) complexes. Differences in chemical shifts between the GSH and GS-hexyl complexes suggest more extensive structural differences between these two enzyme-ligand 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 temperatures in the absence of ligand. A single conformation of alpha2 is induced by the presence of either GSH or GS-hexyl or a reduction in temperature to below 290 K. The large enthalpy of the transition ( approximately 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 to -6 kJ/mol at physiological temperatures (37 degrees C). This order-to-disorder transition contributes substantially to the overall thermodynamics of ligand binding and should be considered in the design of selective inhibitors of class pi glutathione transferases.