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

Abstract

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 , mediumchain-length (MCL) PHAs consist of monomer units of C 6 to C 14 , and SCL-MCL PHAs 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 PHA 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 PHA 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 PHA 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 PHA production.

Comparative metabolomic analysis of Sinorhizobium sp. C4 during the degradation of phenanthrene

“…Limited bioavailability of PAHs and slow transformation to the metabolites in primary metabolism may be the cause of inhibition of PHA biosynthesis during phenanthrene degradation. It is noteworthy that de novo biosynthesis of fatty acid and PHAs shares common intermediates, ketoacyl-acyl-carrierproteins (Nomura et al 2004;Rehm et al 2001). Because of the possible competition between the two metabolic pathways, changes in fatty acid profiles may be interconnected with PHA contents.…”

Advanced functionalization of polyhydroxyalkanoate via the UV-initiated thiol-ene click reaction

“…Copolymerization with various hydroxyalkanoate (HA) units is known to be effective in improving the physical properties and functionalities of PHA, and therefore, the syntheses of various copolymers have been attempted (Dai et al 2008;Fukui et al 1999;Han et al 2009;Hu et al 2011;Matsumoto et al 2011;Matsusaki et al 1998;Nomura et al 2004;Page et al 1992;Shozui et al 2009;Tajima et al 2009;Tajima et al 2012;Tanadchangsaeng et al 2009;Valentin et al 2000;Valentin, Reister and Gruys 2000). However, few studies have been carried out into the introduction of monomer units bearing amino and carboxyl groups, which exhibit high reactivity and are useful for PHA functionalization.…”

“…PHA synthases, which are key enzymes in PHA biosynthesis, catalyze the polymerization of (R)-hydroxyalkanoyl moieties in (R)-hydroxyalkanoyl coenzyme A (HACoA), with the concomitant release of CoA (Rehm 2003;Taguchi and Doi 2004). The…”

Expression of 3-Ketoacyl-Acyl Carrier Protein Reductase ( fabG ) Genes Enhances Production of Polyhydroxyalkanoate Copolymer from Glucose in Recombinant Escherichia coli JM109

“…In our previous study, we demonstrated that coexpression of mutant 3-ketoacyl-acyl carrier protein (ACP) synthase III genes (fabH) with PHA synthase genes (phaC) leads to the production of SCL-MCL PHA in recombinant Escherichia coli grown in the presence of excess glucose (16). That study proposes that substrates for PHA production are derived from the fatty acid biosynthesis pathway (Fig.…”