Mutational analysis of malate pathways in Schizosaccharomyces pombe

Abstract

Schizosaccharomyces pombe is of interest to wine-makers because, unlike the Saccharomyces cerevisiae wine yeasts, it efficiently metabolizes all levels of malic acid found in grape musts. To determine the metabolic pathway for malate metabolism in Sch. pombe, a mutational analysis was performed using a selection system which isolated 154 mutants that were unable to utilize malic acid. The mutants were in three complementation groups which corresponded to alleles and activities for malate dehydrogenase, malic enzyme and malate transport respectively. Under anaerobic conditions Sch. pombe ferments malic acid by using a malate transport, malic enzyme, pyruvate decarboxylase, alcohol dehydrogenase pathway (a malo-ethanolic fermentation) but under aerobic conditions malate is both fermented to ethanol and to a much lesser extent, respired as a TCA cycle intermediate. Alleles were mapped genetically and the locations were confirmed by chromoblotting. Malic enzyme and malate transport alleles were cloned by complementation and the proteins have been purified and partially characterized.