A novel strategy for l-arginine production in engineered Escherichia coli

Abstract Background l -arginine is an important amino acid with applications in diverse industrial and pharmaceutical fields. n -acetylglutamate, synthesized from l -glutamate and acetyl-CoA, is a precursor of the l -arginine biosynthetic branch in microorganisms. The enzyme that produces n -acetylglutamate, n -acetylglutamate synthase, is allosterically inhibited by l -arginine. l -glutamate, as a central metabolite, provides carbon backbone for diverse biological compounds besides l -arginine. When glucose is the sole carbon source, the theoretical maximum carbon yield towards l -arginine is 96.7%, but the experimental highest yield was 51%. The gap of l -arginine yield indicates the regulation complexity of carbon flux and energy during the l -arginine biosynthesis. Besides endogenous biosynthesis, n -acetylglutamate, the key precursor of l -arginine, can be obtained by chemical acylation of l -glutamate with a high yield of 98%. To achieve high-yield production of l -arginine, we demonstrated a novel approach by directly feeding precursor n -acetylglutamate to engineered Escherichia coli . Results We reported a new approach for the high yield of l -arginine production in E. coli. Gene argA encoding n -acetylglutamate synthase was deleted to disable endogenous biosynthesis of n -acetylglutamate. The feasibility of external n -acetylglutamate towards l -arginine was verified via growth assay in argA − strain. To improve l -arginine production, astA encoding arginine n -succinyltransferase, speF encoding ornithine decarboxylase, speB encoding agmatinase, and argR encoding an arginine responsive repressor protein were disrupted. Based on overexpression of argDGI, argCBH operons , encoding enzymes of the l -arginine biosynthetic pathway, ~ 4 g/L l -arginine was produced in shake flask fermentation, resulting in a yield of 0.99 mol l -arginine/mol n -acetylglutamate. This strain was further engineered for the co-production of l -arginine and pyruvate by removing genes adhE, ldhA, poxB, pflB, and aceE, encoding enzymes involved in the conversion and degradation of pyruvate . The resulting strain was shown to produce 4 g/L l -arginine and 11.3 g/L pyruvate in shake flask fermentation. Conclusions Here, we developed a novel approach to avoid the strict regulation of l -arginine on ArgA and overcome the metabolism complexity in the l -arginine biosynthesis pathway. We achieve a high yield of l -arginine production from n -acetylglutamate in E. coli . Co-production pyruvate and l -arginine was used as an example to increase the utilization of input carbon sources.