Metabolically engineered Escherichia coli for efficient production of glycosylated natural products.
Bottom Line: Significant achievements in polyketide gene expression have made Escherichia coli one of the most promising hosts for the heterologous production of pharmacologically important polyketides.However, attempts to produce glycosylated polyketides, by the expression of heterologous sugar pathways, have been hampered until now by the low levels of glycosylated compounds produced by the recombinant hosts.By carrying out metabolic engineering of three endogenous pathways that lead to the synthesis of TDP sugars in E. coli, we have greatly improved the intracellular levels of the common deoxysugar intermediate TDP-4-keto-6-deoxyglucose resulting in increased production of the heterologous sugars TDP-L-mycarose and TDP-D-desosamine, both components of medically important polyketides.
Affiliation: Microbiology Division, IBR (Instituto de Biología Molecular y Celular de Rosario), Consejo Nacional de Investigaciones Científicas y Técnicas, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Rosario, Argentina.Show MeSH
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Mentions: Most of the deoxysugars in natural bioactive compounds belong to the 6‐deoxyhexoses (6DOH) family (Salas and Mendez, 2007). Many of them, like TDP‐l‐mycarose and TDP‐d‐desosamine from the erythromycin and megalomicin biosynthetic pathways, are synthesized through the common precursor TDP‐4‐keto‐6‐deoxyglucose (TKDG). This metabolite is generated from glucose‐1‐phosphate activated to TDP‐d‐glucose by a TDP‐d‐glucose synthase (e.g. RmlA), and further dehydratated to TDP‐4‐keto‐6‐deoxy‐d‐glucose by a TDP‐d‐glucose‐4,6‐dehydratase (e.g. RmlB) (Fig. 1A). In E. coli, TKDG is a precursor of TDP‐4‐acetamido‐4,6‐dideoxy‐d‐galactose (TDP‐d‐Fuc4NAc) (Fig. 1A), a nucleotide‐activated sugar involved in the biosynthesis of the enterobacterial common antigen (ECA), a glycolipid located on the cell surface of all Gram‐negative enteric bacteria (Kajimura et al., 2006). TKDG is also an intermediate in the synthesis of sugars that form the O‐specific polysaccharide chain of the lipopolysaccharide (LPS) described in several E. coli serotypes, like TDP‐l‐rhamnose and TDP‐4‐acetamido‐4,6‐dideoxyglucose (TDP‐d‐Qui4NAc) of the O7 LPS in E. coli VW187 (Marolda et al., 1999; Fig. 1A). The presence of endogenous TKDG‐consuming pathways in E. coli suggests the possibility of a limited availability of this compound for the heterologous biosynthesis of TDP‐l‐mycarose and TDP‐d‐desosamine which, in turn, might result in low levels of glycosylated polyketides in this host.
Affiliation: Microbiology Division, IBR (Instituto de Biología Molecular y Celular de Rosario), Consejo Nacional de Investigaciones Científicas y Técnicas, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Rosario, Argentina.