Metabolic engineering of Corynebacterium glutamicum for the fermentative production of halogenated tryptophan

Halogenated compounds, like 7-chloro-l-tryptophan, are important intermediates or components of bioactive substances relevant for the pharmaceutical, chemical and agrochemical industries. About 20% of all pharmaceutical small molecule drugs and around 30% of all active compounds in agrochemistry are halogenated. Chemical halogenation procedures usually are characterized by the use of hazardous or even highly toxic chemicals. Recently, a biocatalytic process for l-tryptophan halogenation at the gram-scale using FAD-dependent halogenase and NADH-dependent flavin reductase enzymes has been described. Many proteinogenic amino acids are produced by fermentation using Corynebacterium glutamicum. The fermentative production of l-glutamate and l-lysine, for example, is operated at the million-ton scale. However, fermentative production of halogenated amino acids has not yet been described. In this study, fermentative production of the halogenated amino acid 7-chloro-l-tryptophan from sugars, ammonium and chloride salts was achieved. This required metabolic engineering of an l-tryptophan producing C. glutamicum strain for expression of the genes coding for FAD-dependent halogenase RebH and NADH-dependent flavin reductase RebF from Lechevalieria aerocolonigenes. Chlorination of l-tryptophan to 7-chloro-l-tryptophan by recombinant C. glutamicum was improved by optimizing the RBS of rebH. Metabolic engineering enabled production of 7-chloro-l-tryptophan and l-tryptophan from the alternative carbon sources arabinose, glucosamine and xylose.