Kinetics and mechanism of pepsinogen activation.

The spontaneous and pepsin-catalyzed activation of pepsinogen has been observed and analyzed kinetically. At appropriate protein concentrations (1 mg per ml or less), a kinetically first order reaction was observed in the pH range 1 to 3, implying an intramolecular activation mechanism. Substantiation of the first order reaction came from a linear plot of log pepsinogen concentration versus time, with an ordinate intercept at the log of the initial pepsinogen concentration. Moreover a IO-fold dilution in protein concentration did not diminish the half-life of the activation reaction. The first order rate constant of 4.7 per min at 28” was comparable to the k,,$ values observed for peptic hydrolysis of synthetic peptide substrates. At pH 4, we observed kinetics consistent with a mixed reaction mechanism involving the bimolecular reaction of a pepsin molecule and a pepsinogen molecule as well as an intramolecular activation reaction. Below pH 3, this second order process was studied by observing the initial rate of pepsinogen activation in the presence of pepsin. When the pH profiles of the first and second order rate constants were compared, the first order rate constant declined much more rapidly as pH increased than did the second order rate constant. This fact explains the change from a predominantly intermolecular activation mechanism at pH 4 to an intramolecular activation below pH 3.