COMPARISON OF THE DENATURATION BY GUANIDINIUM CHLORIDE BETWEEN ALL OF-THE SITES AND HALE OF-THE-SITES MODIFIED GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE
3-磷酸甘油醛脱氢酶的胍变性——全位与半位修饰酶的比较

Under carefully controlled conditions, yeast GAPDH carboxymethylated at only 2 or all 4 of the subunits can be respectively produced fluorescent NAD derivative at 1 or 2 of the decarboxylated subunits by irradiation in presence of NAD+ (Tsou, et al, Biochem. Soc. Trans. 11, 425-429, 1983). The emission peak at 410nm of the fluorescent NAD derivative introduced at the active site of this enzyme shows both a red shift and a marked decrease in intensity at the same GuHCl concentration required to bring about the inactivation and the initial changes in the intrinsic fluorescence of the holoenzyme. The conformation of modified GAPDH carrying the fluorescent NAD derivative at one subunit is relatively less perturbed than that carrying the derivative at two subunits. The kinetics of this emission change is also biphasic with a fast phase approaching that for the fast phase of inactivation. It appears that treatment by low concentration GuHCl leads both inactivation and perturbation of the active site conformation and these are then followed by unfolding of the molecule as a whole.