Metabolic Control Analysis was applied to calculate the rate-controlling steps of the glycolytic flux of Saccharomyces cerevisiae growing aerobically in chemostat cultures subjected to carbon, nitrogen, or phosphate limitations. The control over the flux ratio at the branch point to ethanol formation and the tricarboxylic acid cycle as well as metabolite concentration control coefficients were determined. Two sorts of models were analyzed, essentially differing in the glucose uptake step which was considered either ATP-and extracellular glucose-dependent (model I) or glucose 6-phosphate-dependent (model II). Kinetic parameters were optimized according to the experimentally determined metabolite concentrations. In both models, glucose uptake was the main rate-controlling step of the glycolytic flux. In model I the control coefficient of the uptake varied between 1.01 and 1.043. The control was shared by ATP-consuming and -producing processes (C-ATPase(J) = -0.080 to -0.0306; C-ADH(J) = -0.0069 to -0.0144; C-TCA(J) = 0.0044 to 0.0144). Meanwhile, in model II of the flux was almost exclusively controlled by the uptake (C-IN(J) = 0.99). Independently of the nutrient limitation, growth rate and type of glucose breakdown metabolism, the rate-controlling steps were the same and only quantitative differences were noted. All metabolite concentrations were controlled by the same rate-controlling steps of the flux, and additionally, by the reaction steps catalyzing the metabolite consumption. The results presented point to the significant role of sugar uptake in yeast catabolism and the usefulness of control analysis to understand metabolic flux as applied to biotransformation processes.