Effects of pH in rapid-equilibrium enzyme kinetics

The effects of pH on the rates of enzyme-catalyzed reactions are very important because they yield information on the pKs of acidic groups in the enzymatic site and the various enzyme-substrate complexes. But many enzyme-catalyzed reactions produce or consume hydrogen ions in a way that cannot be explained with pKs. These pH effects extend over the whole pH range of interest. In investigating these effects, the rapid-equilibrium assumption is especially useful because a large number of chemical reactions have to be taken into account. In these calculations, all of the reactions up to the rate-determining reaction are treated with biochemical thermodynamics. Kinetic studies make it possible to determine the number of hydrogen ions consumed in the rate-determining reaction, a number that can be in the range of 0-8. It is shown that the experimental limiting velocity of the forward reaction V(fexp) is equal to 10(npH)V(f), where n is a negative integer and Vf varies with pH in the way determined by the pKs of the enzyme-substrate complex that reacts in the rate-determining reaction. A computer program for the initial reaction velocity makes it possible to investigate the rapid-equilibrium kinetics of enzymatic mechanisms that involve the consumption of hydrogen ions.