ENGINEERED PORCINE PEPSINOGEN EXHIBITS DOMINANT UNIMOLECULAR ACTIVATION

An engineered pepsinogen, which was a fusion protein of thioredoxin an d pepsinogen, exhibited dominant self-activation (unimolecular reactio n; intramolecular activation) in contrast to recombinant pepsinogen wh ich exhibited both unimolecular and bimolecular reactions (intermolecu lar activation mediated by pepsin released during activation). At pH v alues of 1.1, 2.0, and 3.0, activation curves for the engineered pepsi nogen were hyperbolic rather than sigmoidal, indicating that self-acti vation was the dominant activation mechanism in comparison to the slow er bimolecular activation. To confirm which activation mechanism was d ominant, an equal mole of pepsin was added to accelerate the bimolecul ar reaction during activation. The addition of exogenous pepsin did no t affect the activation rate of the engineered pepsinogen but accelera ted pepsinogen activation through the bimolecular reaction. The above results indicated that the engineered pepsinogen exhibited, primarily, a self-activation mechanism and that bimolecular activation was negli gible. (C) 1997 Academic Press.