A thermal-variation method for analysing the rate constants of the Michaelis--Menten mechanism.

By analysing the variations of saturation velocity and Michaelis constant with temperature and invoking the mathematical constraint represented by the Arrhenius equation, it becomes possible to estimate k+2 and indistinguishably k+1 and k-1 for the Michaelis--Menten mechanism of one-substrate enzyme reactions. Distinction between k+1 and k-1 may be obtained through the determination of isotopic rate effects. This procedure thus provides a basis for evaluating all three rate constants of the one-substrate mechanism, and disproves the suggestion that k+1 and k-1 are intrinsically unobtainable from steady-state kinetic measurements.