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

Abstract

AbstractBackgroundl-arginine is an important amino acid with applications in diverse industrial and pharmaceutical fields.n-acetylglutamate, synthesized froml-glutamate and acetyl-CoA, is a precursor of thel-arginine biosynthetic branch in microorganisms. The enzyme that producesn-acetylglutamate,n-acetylglutamate synthase, is allosterically inhibited byl-arginine.l-glutamate, as a central metabolite, provides carbon backbone for diverse biological compounds besidesl-arginine. When glucose is the sole carbon source, the theoretical maximum carbon yield towardsl-arginine is 96.7%, but the experimental highest yield was 51%. The gap ofl-arginine yield indicates the regulation complexity of carbon flux and energy during thel-arginine biosynthesis. Besides endogenous biosynthesis,n-acetylglutamate, the key precursor ofl-arginine, can be obtained by chemical acylation ofl-glutamate with a high yield of 98%. To achieve high-yield production ofl-arginine, we demonstrated a novel approach by directly feeding precursorn-acetylglutamate to engineeredEscherichia coli.ResultsWe reported a new approach for the high yield ofl-arginine production inE. coli.GeneargAencodingn-acetylglutamate synthase was deleted to disable endogenous biosynthesis ofn-acetylglutamate. The feasibility of externaln-acetylglutamate towardsl-arginine was verified via growth assay inargA−strain. To improvel-arginine production,astAencoding argininen-succinyltransferase,speFencoding ornithine decarboxylase,speBencoding agmatinase, andargRencoding an arginine responsive repressor protein were disrupted. Based on overexpression ofargDGI, argCBHoperons,encoding enzymes of thel-arginine biosynthetic pathway, ~ 4 g/Ll-arginine was produced in shake flask fermentation, resulting in a yield of 0.99 moll-arginine/moln-acetylglutamate. This strain was further engineered for the co-production ofl-arginine and pyruvate by removing genesadhE, ldhA, poxB, pflB,andaceE,encoding enzymes involved in the conversion and degradation of pyruvate.The resulting strain was shown to produce 4 g/Ll-arginine and 11.3 g/L pyruvate in shake flask fermentation.ConclusionsHere, we developed a novel approach to avoid the strict regulation ofl-arginine on ArgA and overcome the metabolism complexity in thel-arginine biosynthesis pathway. We achieve a high yield ofl-arginine production fromn-acetylglutamate inE. coli. Co-production pyruvate andl-arginine was used as an example to increase the utilization of input carbon sources.