BINDING IN THE TRANSITION-STATE FOR UNFOLDING OF ADENOSINE-DEAMINASE

Inhibitors protect calf intestinal adenosine deaminase from reversible denaturation by guanidine hydrochloride, and from heat inactivation. In the presence of saturating concentrations of ligands, the first-ord er rate constant for reversible denaturation of this enzyme in 3.0 M g uanidine hydrochloride is reduced 6-fold by pteridine or 6-dimethylami nopurine ribonucleoside, and 60-fold by purine ribonucleoside; these c ompounds provide similar protection against thermal inactivation at 70 degrees C. These protective effects indicate that in the transition s tate for denaturation by heat or guanidine hydrochloride, the enzyme r etains much of its native structure, exhibiting 60-70% of the free ene rgy of association with these ligands that was present in the native e nzyme in the ground state. However, neither these ligands nor the extr emely powerful inhibitor 2'-deoxycoformycin affect the rate of refoldi ng of enzyme denatured in guanidine hydrochloride. (C) 1994 Academic P ress, Inc.