Viscosimetric Determination of Cellulase Activity in the Intestine of the Sea Urchin: Reaction Mechanism and Equilibrium Constant for Cellulase Stabilization with Calcium.

Abstract

Intestines of the sea urchin Paracentrotus lividus contain a cellulase which is stabilized by calcium. The apparent rate constant k for the inactivation of the enzyme was determined at various calcium ion activities in a buffer solution in which the buffering substances did not react with calcium ions. The rate constant k[mu] for the unstabilized enzyme and the rate constant ks for the stabilized enzyme were calculated. The logarithm of (k[long dash]ks)/(ku[long dash]k) was plotted versus the logarithm of the calcium ion activity, and a straight line was fitted to the points. The inclination of such a line gives the number n of calcium ions reacting with each of those groups of the enzyme which have an influence upon its stability, and the intersection of the line with the axis of abscissas gives the value of the logarithm of the equilibrium constant K for the stabilization of the enzyme, divided by n. In the stabilization of the cellulase studied, each of its chemical groups which have an influence on its stability react with one calcium ion. The equilibrium constant for this stabilizing reaction is 0.0009 mole/liter. The optimum pH value for the stability of this cellulase is about 6.8 and is not significantly affected by the calcium ion activity. The rate constant for the inactivation of the calcium-free cellulase is about 4 times as large as that of its calcium complex. The optimum pH value for activity is about 6.1.