MECHANISM AND ION-DEPENDENCE OF IN-VITRO AUTOACTIVATION OF YEAST PROTEINASE-A - POSSIBLE IMPLICATIONS FOR COMPARTMENTALIZED ACTIVATION IN-VIVO

Yeast proteinase A is synthesized as a zymogen which transits through the endoplasmic reticulum, the Golgi complex and the endosome to the v acuole. On arrival in the vacuole, activation takes place. It has prev iously been found that proteinase A can activate autocatalytically; ho wever, the propeptide of proteinase A shows essentially no similarity to other known aspartic proteinase propeptides. To understand why prot einase A activation occurs rapidly in the vacuole but not at all in ea rlier compartments, we have purified the zymogen and investigated the conditions that trigger autoactivation and the mechanism of autoactiva tion. Autoactivation was triggered by acidic pH and its rate increased with increasing ionic strength. Kinetic evidence indicates that autoa ctivation mainly occurs via a bimolecular product-catalysed mechanism in which an active proteinase A molecule activates a zymogen molecule. Both the pH-and ionic- strength-dependence and the predominance of a product-catalysed mechanism are well adapted to the situation in vivo, since slow activation in the absence of active proteinase A helps to prevent activation in prevacuolar compartments, whereas, on delivery t o the vacuole, lower pH, higher ionic strength and the presence of alr eady active proteinases ensure rapid activation. Product-catalysed aut oactivation may be a general mechanism by which cells ensure autoactiv ation of intracellular enzymes to be both rapid and compartmentalized.