QUANTITATIVE-ANALYSIS OF SCHIZOPHYLLUM-COMMUNE METALLOPROTEASE SCPRB ACTIVITY IN SDS-GELATIN PAGE REVEALS DIFFERENTIAL MYCELIAL LOCALIZATION OF NITROGEN LIMITATION-INDUCED AUTOLYSIS

The basidiomycete Schizophyllum commune produces a variety of proteoly tic enzymes. A number of these, detected in native gelatin-containing polyacrylamide gels, have their activities increased during nitrogen-l imited growth of the mycelium. ScPrB, a metallo-endoprotease, appears to have the greatest nitrogen stress-induced increase in activity of a ll of these enzymes. Quantifying ScPrB has proven difficult because no artificial chromogenic substrate has been found. In addition, it is p oorly resolved from another highly active protease, ScPrA, in native g elatin-containing gels. We have developed a method using SDS gelatin-c ontaining polyacrylamide gels for resolving ScPrB from ScPrA and for q uantifying its activity by densitometry. This method was used to asses s the intramycelial location of ScPrB induction after the transfer of exponentially growing colonies to nitrogen deprivation conditions. By all analyses (proportional, normalized to fresh weight, and normalized to protein), the increase in ScPrB activity was found to occur exclus ively in midsections of the growing mycelium, whereas ScPrB activity w as found to be decreased or unchanged in the centers of colonies and i n colony margins. This implies that proteolysis mediated by ScPrB may supply translocatable amino acids only from the region directly behind the growing hyphal apices.