This paper is concerned with the control of glycolysis in nongrowing Streptococcus lactis 7962. Changes were measured in the concentrations of glycolytic intermediates, intracellular inorganic phosphate (Pi), and adenine nucleotides following addition of glucose to cells that were in a starved condition. We find that intracellular Pi is a major factor in the control of glycolysis. In starved cells, the intracellular Pi concentration is high, greater than 40 mM. The large phosphoenolpyruvate pool that exists in starved cells can be explained as a result of inhibition of pyruvate kinase by the high concentration of Pi. On the other hand, in cells that are metabolizing glucose at a steady state rate, the cellular Pi concentration is low and pyruvate kinase is active. Upon depletion of glucose from the medium, the metabolite concentrations return to the values originally found in the starved state. This glucose depletion raises the intracellular Pi which again leads to inhibition of pyruvate kinase and the consequent buildup of the P-enolpyruvate pool.