Intracellular acetyl-CoA depletion by the cat gene is responsible for growth inhibition of Escherichia coli C584 on M9 minimal media

Abstract

Two closely related strains of Escherichia coli, C584 and DH5 α, differ in their ability to grow on M9 minimal media supplemented with glycerol and thiamine. While the DH5 α strain is able to grow in the nutrient-limited medium, the identical C584 strain containing the pMOB3 plasmid fails to do so. The chloramphenicol resistance cat gene encoded on the pMOB3 plasmid has been implicated in this growth inhibition. The proposed mechanism of the inhibition is by the action of the cat gene product chloramphenicol acetyl transferase (CAT). CAT utilizes acetyl-CoA, depleting the cell of an important precursor metabolite, resulting in cell growth arrest. The purpose of our study was to determine whether acetyl-CoA depletion occurs in the C584 strain incubated in M9 minimal media, and if so, whether sodium acetate, an extracellular source of acetyl-CoA, can replenish acetyl-CoA levels and allow for the growth of E. coli C584 in M9 minimal media. Our results suggest that intracellular acetyl-CoA levels decrease in E. coli C584 grown in M9 minimal media and that sodium acetate supplementation restores these levels to sufficient levels, enabling E. coli C584 growth.