Multiple Strategies for Metabolic Engineering of Escherichia coli for Efficient Production of Coenzyme Q10

Abstract

Abstract: Escherichia coli BW25113 was metabolically engineered for CoQ₁₀ production by replacing ispB with ddsA from Gluconobacter suboxydans.Effects of precursor balance and reduced nicotinamide-adenine dinucleotide phosphate (NADPH) availability on CoQ₁₀ production in E.coli were investigated.The knockout of pykFA along with pck overexpression could maintain a balance between glyceraldehyde 3-phosphate and pyruvate,increasing CoQ₁₀ production.Replacement of native NAD-dependent gapA with NADP-dependent gapC from Clostridium acetobutylicum,together with the overexpression of gapC,could increase NADPH availability and then enhanced CoQ₁₀ production.Three effects,overexpressions of various genes in CoQ biosynthesis and central metabolism,different vectors and culture conditions on CoQ₁₀ production in E.coli,were all investigated.The investigation of different vectors indicated that low copy number vector may be more beneficial for CoQ₁₀ production in E.coli.The recombinant E.coli (△ispB::ddsA,△pykFA and △gapA::gapC),harboring the two plasmids encoding pck,dxs,idi and ubiCA genes under the control of P_T5 on pQE30,ispA,ddsA from Gluconobacter suboxydans and gapC from Clostridium acetobutylicum under the control of PBAD on pBAD33,could produce CoQ₁₀ up to 3.24 mg·g^-1 dry cell mass simply by changing medium from M9YG to SOB with phosphate salt and initial culture pH from 7.0 to 5.5.The yield is unprecedented and 1.33 times of the highest production so far in E.coli.

Key words: coenzyme Q₁₀, Escherichia coli, gene replacement, NADPH availability, precursor balance

摘要： Escherichia coli BW25113 was metabolically engineered for CoQ₁₀ production by replacing ispB with ddsA from Gluconobacter suboxydans.Effects of precursor balance and reduced nicotinamide-adenine dinucleotide phosphate (NADPH) availability on CoQ₁₀ production in E.coli were investigated.The knockout of pykFA along with pck overexpression could maintain a balance between glyceraldehyde 3-phosphate and pyruvate,increasing CoQ₁₀ production.Replacement of native NAD-dependent gapA with NADP-dependent gapC from Clostridium acetobutylicum,together with the overexpression of gapC,could increase NADPH availability and then enhanced CoQ₁₀ production.Three effects,overexpressions of various genes in CoQ biosynthesis and central metabolism,different vectors and culture conditions on CoQ₁₀ production in E.coli,were all investigated.The investigation of different vectors indicated that low copy number vector may be more beneficial for CoQ₁₀ production in E.coli.The recombinant E.coli (△ispB::ddsA,△pykFA and △gapA::gapC),harboring the two plasmids encoding pck,dxs,idi and ubiCA genes under the control of P_T5 on pQE30,ispA,ddsA from Gluconobacter suboxydans and gapC from Clostridium acetobutylicum under the control of PBAD on pBAD33,could produce CoQ₁₀ up to 3.24 mg·g^-1 dry cell mass simply by changing medium from M9YG to SOB with phosphate salt and initial culture pH from 7.0 to 5.5.The yield is unprecedented and 1.33 times of the highest production so far in E.coli.

关键词: coenzyme Q₁₀, Escherichia coli, gene replacement, NADPH availability, precursor balance

HUANG Mingtao, WANG Yue, LIU Jianzhong, MAO Zongwan. Multiple Strategies for Metabolic Engineering of Escherichia coli for Efficient Production of Coenzyme Q₁₀[J]. , 2011, 19(2): 316-326.

黄明涛, 王玥, 刘建忠, 毛宗万. Multiple Strategies for Metabolic Engineering of Escherichia coli for Efficient Production of Coenzyme Q₁₀[J]. , 2011, 19(2): 316-326.

References

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Multiple Strategies for Metabolic Engineering of Escherichia coli for Efficient Production of Coenzyme Q₁₀

Multiple Strategies for Metabolic Engineering of Escherichia coli for Efficient Production of Coenzyme Q₁₀

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