Studies on a Proteinase B Mutant of Yeast

Yeast mutants lacking proteinase B activity have been isolated [Wolf, D. H. and Ehmann, C. (1978) FEBS Lett. 92, 121–124]. One of these mutants (HP232) is characterized in detail. Absence of the vacuolar localized enzyme is confirmed by checking for proteinase B activity in isolated mutant vacuoles. Defective proteinase B activity segregates 2:2 in meiotic tetrads. The mutation is shown to be recessive. Mutant proteinase B activity is not only absent against the synthetic substrate, Azocoll, but also against the physiological substrates pre‐chitin synthetase, cytoplasmic malate dehydrogenase and fructose‐1,6‐bisphosphatase. The mutant shows normal vegetative growth, a phenomenon not consistent with the idea that proteinase B might be the activating principle of chitin synthetase zymogen in vivo. Fluorescence microscopy shows normal chitin insertion. Enzymes underlying carbon‐catabolite inactivation in wild‐type cells (a mechanism proposed to be possibly triggered by proteinase B) such as cytoplasmic malate dehydrogenase, fructose‐1,6‐bisphosphatase, phosphoenolpyruvate carboxykinase and isocitrate lyase, are inactivated also in the mutant. NADP‐dependent glutamate dehydrogenase, which is found to be inactivated in glucose‐starved wild‐type cells, proceeds normally in the mutant. Mutant cells show more than 40% reduced protein degradation under starvation conditions. Sporulating diploids, homozygous for proteinase B absence, also exhibit an approximately 40% reduced protein degradation as compared to homozygous wild‐type diploids or diploids heterozygous for the mutant gene. The time of the appearance of the first ascospores of diploid cells, homozygous for proteinase B deficiency, is delayed about 50% and sporulation frequency is reduced to about the same extent as compared to homozygous wild‐type diploids or diploids heterozygous for the mutant gene.