title: Metabolic engineering of Corynebacterium glutamicum to produce GDP-L-fucose from glucose and mannose.
abstract: Wild-type Corynebacterium glutamicum was metabolically engineered to convert glucose and mannose into guanosine 5'-diphosphate (GDP)-L-fucose, a precursor of fucosyl-oligosaccharides, which are involved in various biological and pathological functions. This was done by introducing the gmd and wcaG genes of Escherichia coli encoding GDP-D-mannose-4,6-dehydratase and GDP-4-keto-6-deoxy-D-mannose-3,5-epimerase-4-reductase, respectively, which are known as key enzymes in the production of GDP-L-fucose from GDP-D-mannose. Coexpression of the genes allowed the recombinant C. glutamicum cells to produce GDP-L-fucose in a minimal medium containing glucose and mannose as carbon sources. The specific product formation rate was much higher during growth on mannose than on glucose. In addition, the specific product formation rate was further increased by coexpressing the endogenous phosphomanno-mutase gene (manB) and GTP-mannose-1-phosphate guanylyl-transferase gene (manC), which are involved in the conversion of mannose-6-phosphate into GDP-D-mannose. However, the overexpression of manA encoding mannose-6-phosphate isomerase, catalyzing interconversion of mannose-6-phosphate and fructose-6-phosphate showed a negative effect on formation of the target product. Overall, coexpression of gmd, wcaG, manB and manC in C. glutamicum enabled production of GDP-L-fucose at the specific rate of 0.11 mg g cell(-1) h(-1). The specific GDP-L-fucose content reached 5.5 mg g cell(-1), which is a 2.4-fold higher than that of the recombinant E. coli overexpressing gmd, wcaG, manB and manC under comparable conditions. Well-established metabolic engineering tools may permit optimization of the carbon and cofactor metabolisms of C. glutamicum to further improve their production capacity.
authors: Department of Agricultural Biotechnology, Center for Food and Bioconvergence, Seoul National University, Seoul, 151-921, Republic of Korea.
authors: Young-Wook
authors: YW
authors: Chin
authors: Jin-Byung
authors: JB
authors: Park
authors: Yong-Cheol
authors: YC
authors: Park
authors: Kyoung Heon
authors: KH
authors: Kim
authors: Jin-Ho
authors: JH
authors: Seo
mesh_terms: D002238:Carbohydrate Epimerases / Q000096:biosynthesis / Q000235:genetics; D048230:Corynebacterium glutamicum / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D029968:Escherichia coli Proteins / Q000096:biosynthesis / Q000235:genetics; D015870:Gene Expression; D005947:Glucose / Q000378:metabolism* / Q000494:pharmacology; D006154:Guanosine Diphosphate Fucose / Q000096:biosynthesis* / Q000235:genetics; D006836:Hydro-Lyases / Q000096:biosynthesis / Q000235:genetics; D007658:Ketone Oxidoreductases / Q000096:biosynthesis / Q000235:genetics; D008358:Mannose / Q000378:metabolism* / Q000494:pharmacology; D060847:Metabolic Engineering / Q000379:methods; D009097:Multienzyme Complexes / Q000096:biosynthesis / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D013549:Sweetening Agents / Q000378:metabolism / Q000494:pharmacology
chemical_list: D029968:Escherichia coli Proteins; D009097:Multienzyme Complexes; D011994:Recombinant Proteins; D013549:Sweetening Agents; C110682:wcaG protein, E coli; D006154:Guanosine Diphosphate Fucose; D007658:Ketone Oxidoreductases; D006836:Hydro-Lyases; C044971:GDPmannose 4,6-dehydratase; D002238:Carbohydrate Epimerases; D005947:Glucose; D008358:Mannose