The Effect of Temperature on the Catalytic Activity of Bovine Carbonic Anhydrase

Abstract

The two high activity forms of carbonic anhydrase which have been extensively studied, electrophoretic component C of the enzyme from human red cells and electrophoretic component B from bovine red cells, both show remarkably high catalytic activity. Under optimal conditions and at 25 C each molecule of enzyme catalyses the hydration of about 10 molecules of CO₂ per second. This observation which is well established, (Kernohan, 1965; Khalifah, 1971) conflicts with a prediction by Eigen and Hammes (1963) that enzymes having acid-base catalytic mechanisms should have an upper limit of about 10⁵ per second to their catalytic rate. This prediction comes directly from the measurement of the rate constants for the combination of protons with various bases in aqueous solution. These rate constants are found to have an upper limit of about 10¹¹M^-1sec^-1 which corresponds to the collision frequency of the two reactants. For an acid with pK 6 it can be calculated that the rate constant for its dissociation of a proton cannot exceed 10⁵ per second. For acids with higher pK values or whose conjugate bases have lower rate constants for proton addition, the rate constants for proton dissociation will be correspondingly lower. If such a group is successively protonated and then deprotonated during the catalytic action of an enzyme then the rate of the deprotonation reaction will limit the overall catalytic rate which can be achieved.