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 vacuole. On arrival in the vacuole, activation takes place. It has previously been found that proteinase A can activate autocatalytically; however, the propeptide of proteinase A shows essentially no similarity to other known aspartic proteinase propeptides. To understand why proteinase A activation occurs rapidly in the vacuole but not at all in earlier compartments, we have purified the zymogen and investigated the conditions that trigger autoactivation and the mechanism of autoactivation. Autoactivation was triggered by acidic pH and its rate increased with increasing ionic strength. Kinetic evidence indicates that autoactivation 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 to the vacuole, lower pH, higher ionic strength and the presence of already active proteinases ensure rapid activation. Product-catalysed autoactivation may be a general mechanism by which cells ensure autoactivation of intracellular enzymes to be both rapid and compartmentalized.
Van Den Hazel
H
H
Department of Yeast Genetics, Carlsberg Laboratory, Copenhagen, Denmark.
Wolff
A M
AM
Kielland-Brandt
M C
MC
Winther
J R
JR
D016282:Aspartic Acid Endopeptidases / Q000302:isolation & purification / Q000378:metabolism* / Q000502:physiology; D002451:Cell Compartmentation; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000302:isolation & purification / Q000378:metabolism / Q000502:physiology; D006863:Hydrogen-Ion Concentration; D009994:Osmolar Concentration; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000254:growth & development / Q000502:physiology; D014617:Vacuoles / Q000201:enzymology
D004792:Enzyme Precursors; D016282:Aspartic Acid Endopeptidases; C022442:aspergillopepsin II