Comparison of kinetic and optical changes during guanidine denaturation of intestinal alkaline phosphatase.

The modifications of the activity of calf intestinal Alkaline Phosphatase treated with moderate amounts of guanidinium chloride are compared with the conformational changes observed by ultraviolet absorbance and intrinsic fluorescence. The time course of catalytic and optical properties of the treated enzyme develops through two distinct steps: an instantaneous and a time-dependent one. The immediate effect of guanidine is to lower emission yield, to shift the emission maximum of the enzyme to longer wavelengths and to enhance the absorbance of the protein. The rapid conformational transition determines a paradoxical activation at low effector concentration (below 0.88 M) and an inhibition at higher amounts. The following marked decay of enzyme activity with time is related to spectroscopically detectable changes. Temperature influences both kinetic and structural aspects of the process and facilitates guanidine action.