COMPARISON OF INACTIVATION AND UNFOLDING OF YEAST ALCOHOL-DEHYDROGENASE DURING THERMAL-DENATURATION

It has been reported that inactivation occurs before noticeable confor mational change can be detected during denaturation of creatine kinase (ATP:creatine N-phosphotransferase, EC 2.7.3.2) and other enzymes by guanidinium chloride or urea. It has therefore been suggested that enz yme active sites may display more conformational flexibility than the enzyme molecules as a whole. The present paper compares the inactivati on and unfolding of yeast alcohol dehydrogenase during thermal denatur ation. Under identical conditions, inactivation takes place before not iceable conformational changes. Kinetics of unfolding can be resolved into two phases. For a given temperature, the fast phase rates are abo ut one order of magnitude slower than the inactivation rates of the fr ee enzyme and approximately the same magnitude as the inactivation rat es of enzyme-substrate complexes. This is in general accord with the s uggestion made previously by Tsou, indicating that the active sites of metal enzymes are situated in a region more flexible than the molecul es as a whole. (C) 1997 Elsevier Science Ltd.