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Title: BIOSYNTHESIS OF MEDIUM-CHAIN-LENGTH POLY(HYDROXYALKANOATES) BY ALTERED COMPOSITION BY MUTANT HYBRID PHA SYNTHASES

Author
item Solaiman, Daniel

Submitted to: Journal of Industrial Microbiology and Biotechnology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/11/2003
Publication Date: 5/1/2003
Citation: Solaiman, D. 2003. Biosynthesis of medium-chain-length poly(hydroxyalkanoates) by altered composition by mutant hybrid PHA Synthases. Journal of Industrial Microbiology and Biotechnology. 30:322-326.

Interpretive Summary: Poly(hydroxyalkanoates) (PHAs) are biopolyesters produced by many bacteria. When isolated from different bacteria, these biopolymers have properties similar to those of hard plastics, stretchy elastomers, or sticky adhesives. These bacterial polymers are biodegradable, and can be produced from renewable resources by feeding the cells with agricultural fats, oils, and byproducts. As such, PHAs potentially are environmentally beneficial substitutes for the fossil fuel-based polymers currently used in many consumer products. To expand the usefulness of these materials, PHAs with new compositions and improved performance properties are desired. In this report, the researcher showed a method to make new genes from the ones naturally found in a bacterium (called Pseudomonas resinovorans) that produces the stretch elastomeric and sticky types of PHAs. When activated in another bacterium (called E. coli), some of these newly generated genes direct the production of polymers with a different composition (or make-up). This study thus demonstrated the validity of the method of mixing naturally existing genes to generate new ones capable of producing PHA with an altered specification. Further research should help produce novel polymers that fit important applications, eventually leading to the manufacture of biobased end-user products that will benefit the consumer.

Technical Abstract: Pseudomonas resinovorans harbors two isogenic poly(hydroxyalkanoates) (PHA) synthase genes (phaC1Pre and phaC2Pre) responsible for the production of intracellular medium-chain-length (mcl-) PHA. Sequence analysis showed that the putative gene-products of these genes contain a conserved alpha/beta-hydrolase fold in the carboxy-terminal half of the proteins. Hybrid genes pha7 and pha8 were constructed by exchanging the 3'-terminal, alpha/beta-hydrolase-fold coding portions of the phaC1Pre and phaC2Pre. When grown with decanoate as carbon source, the pha7- or pha8-transformed E. coli LS1298 produced PHAs containing 73-75% beta-hydroxydecanoate (beta-HD) and 25-27% beta-hydroxyoctanoate (beta-HO). Deletion mutants, delta-pha7 and delta-pha8, were isolated during the PCR-based construction of pha7 and pha8, respectively. Cells harboring these mutants produced PHAs containing 55-60 mol% beta-HD and 40-45 mol% beta-HO. These results demonstrated the feasibility of generating active hybrid mcl-PHA synthase genes and their mutants with the potential of producing polymers having a varied repeat-unit composition.