Coexpression of genetically engineered 3-ketoacyl-ACP synthase III (fabH) and polyhydroxyalkanoate synthase (phaC) genes leads to short-chain-length-medium-chain-length polyhydroxyalkanoate copolymer production from glucose in Escherichia coli JM109
Coexpression of genetically engineered 3-ketoacyl-ACP synthase III (fabH) and polyhydroxyalkanoate synthase (phaC) genes leads to short-chain-length-medium-chain-length polyhydroxyalkanoate copolymer production from glucose in Escherichia coli JM109
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DOI：
10.1128/aem.70.2.999-1007.2004
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发表时间：
2004-02-01
影响因子：
4.4
通讯作者：
Doi, Y
中科院分区：
文献类型：
--
作者：
Nomura, CT;Taguchi, K;Doi, Y
根据聚合物中单体的大小，聚羟基烷酸酯(PHAs)可分为三种主要类型。短链(SCL)PHA由C-3至C-5的单体单元组成，中链(MCL)PHA由C-6至C-14的单体单元组成，而SCL-MCL PRA由大小从C-4至C-14的单体组成。虽然以前使用重组大肠杆菌的研究表明，SCL或MCL PHA聚合物都可以从葡萄糖中生成，但本研究首次证明了在重组大肠杆菌中可以由葡萄糖生成SCL-MCL PHA共聚物。通过对大肠杆菌3-酮酰基载体蛋白合成酶III基因(FabH)第87位编码氨基酸的密码子进行饱和点突变，将其与含有猪气单胞菌PrA合成酶基因(Phac)的Papac和含有假单胞菌PrA合成酶基因的pPPAC共转化。对菌株61-3的PHA合成酶基因(PhaC1)进行了分析，并对这些菌株从葡萄糖中积累PrA的能力进行了评价。研究发现，几个突变的FabH基因的过表达使重组大肠杆菌能够诱导C-4到C-10单体的产生，从而产生含有SCL和MCL单元的不寻常的PRA共聚物。结果表明，PHA共聚物的组成可能受单体供给酶的控制，进一步强化了脂肪酸生物合成可用于生产PRA的单体供应的观点。
Polyhydroxyalkanoates (PHAs) can be divided into three main types based on the sizes of the monomers incorporated into the polymer. Short-chain-length (SCL) PHAs consist of monomer units of C-3 to C-5, medium-chain-length (MCL) PHAs consist of monomer units of C-6 to C-14, and SCL-MCL PRAs consist of monomers ranging in size from C-4 to C-14. Although previous studies using recombinant Escherichia coli have shown that either SCL or MCL PHA polymers could be produced from glucose, this study presents the first evidence that an SCL-MCL PHA copolymer can be made from glucose in recombinant E. coli. The 3-ketoacyl-acyl carrier protein synthase III gene (fabH) from E. coli was modified by saturation point mutagenesis at the codon encoding amino acid 87 of the FabH protein sequence, and the resulting plasmids were cotransformed with either the pAPAC plasmid, which harbors the Aeromonas caviae PRA synthase gene (phaC), or the pPPAC plasmid, which harbors the Pseudomonas sp. strain 61-3 PHA synthase gene (phaC1), and the abilities of these strains to accumulate PRA from glucose were assessed. It was found that overexpression of several of the mutant fabH genes enabled recombinant E. coli to induce the production of monomers of C-4 to C-10 and subsequently to produce unusual PRA copolymers containing SCL and MCL units. The results indicate that the composition of PHA copolymers may be controlled by the monomer-supplying enzyme and further reinforce the idea that fatty acid biosynthesis may be used to supply monomers for PRA production.