When deprived of iron, Saccharomyces cerevisiae rearranges its metabolic flux towards increased glycerol production. This work examines the role and regulation of GPD1 and GPD2, encoding two isoforms of glycerol 3-phosphate dehydrogenase, in glycerol production during iron starvation. The two genes respond differently on transfer of cells to iron-limited conditions. Whereas the expression of GPD2 increases about 3-fold, that of GPD1 does not exhibit significant changes, Deletion of either GPD1 or GPD2 alters the capacity for glycerol production during iron-limited as well as iron sufficient conditions. However, loss of function of either gene does not seem to provoke compensatory flux via the other gene product. As judged from the glycerol production, the amount produced by each single mutant adds approximately up to the level produced by the parental strain, In agreement with the pattern of expression of GPD2, this gene product was estimated to account for the bulk of the glycerol production (about 60%,) during iron-limited conditions. The strong growth inhibition caused by iron starvation was reversed by the addition of iron also for a gpd1 Delta gpd2 Delta double deletion mutant, which is unable to produce any detectable glycerol. (C) 1999 Federation of European Biochemical Societies.