Accumulation of K+ which is lost during repeated washings stimulates anaerobic glycolysis in E. coli. The gramycidin D-induced outflow of K+ from the bacteria inhibits the glycolysis. An addition of KCl to the cells treated with gramycidin D increases the rate of glycolysis. The concentration dependence of the effectiveness of glycolysis of gramycidin D-treated E. coli cells on KCl concentration has a maximum 100 mM. A direct involvement of K+ in the regulation of anaerobic glycolysis of E. coli is proposed.