Genetic manipulation of a primary metabolic pathway for l-ornithine production in Escherichia coli

Abstract

Metabolic engineering has been used to improve l-ornithine biosynthesis in Escherichia coli W3110. l-Ornithine production increased from 0.3 to 3.2 mg/g (dry cell weight) when the primary l-ornithine biosynthetic pathway was optimized by disrupting the pathway transcription repressor, thereby increasing the expression of the genes involved in the pathway, and by preventing conversion of l-ornithine into citrulline. When a feedback-resistant N-acetylglutamate synthetase gene (argA214) was placed under the control of the arabinose-inducible promoter, either in the chromosome or on a multicopy plasmid in the cell, the combination of overexpression of argA214 with an argF argI argR triple knockout mutation had an additive effect on l-ornithine production but only when exogenous glutamate was present. When speF (which encodes ornithine decarboxylase) and proB (which encodes γ-glutamyl kinase) were inactivated to prevent the conversion of l-ornithine to putrescine and to block the biosynthesis of a side branch of l-ornithine, respectively, l-ornithine production was further enhanced approxi. 140% from 5.5 to 13.2 mg/g (dry cell weight).