AbstractAn E. coli strain in which all known pathways of threonine catabolism were inactivated (Δtdh, ΔltaE, ΔilvA, ΔtdcB, ΔyiaY) has been constructed. It was shown that the expression of heterologous citramalate synthase from Leptospira interrogans in an E. coli strain carrying the ΔilvA deletion can serve as an alternative pathway for isoleucine synthesis. It was observed that cimA overexpression has a negative effect on threonine production. We developed a system for regulated gene expression based on the inducible promoter PLtetO and TetR, a repressor of the tetracycline operon. The threonine-producing strain B-1201, in which the cimA gene is expressed under the control of the regulated promoter, was constructed. When the B-1201 strain was cultivated in a fermenter, a correlation was established between the threonine productivity and the expression level of the cimA gene, and the optimal inductor content for the maximum threonine accumulation was also determined.