Escherichia coli excretes acetate during aerobic growth on LB broth containing glucose and growth ceases before depletion of glucose because of the low pH caused by the accumulation of acetate. It has been known that the acetate accumulation is reduced even when E. coli is grown in the presence of high concentration of glucose if Mlc is overexpressed. The intracellular concentration of Mlc is very low in E. coli because of autoregulation and a low efficiency of mlc translation. We constructed various mutants that can express higher levels of Mlc using site-directed mutagenesis and one of the Mlc-overproducing mutant showed reduced glucose consumption rate and low production of acetate. The mutant showed higher foreign gene expression level than that of its parental strain in the presence of glucose. These results suggest that the Mlc overproducing E. coli strain having an improved ability of glucose utilization can be a better host for high-level production of useful recombinant proteins.