Characterization Of A Highly Active Polyhydroxyalkanoate Synthase
| dc.contributor.author | Chuah, Jo-Ann | |
| dc.date.accessioned | 2019-06-18T06:14:49Z | |
| dc.date.available | 2019-06-18T06:14:49Z | |
| dc.date.issued | 2012-06 | |
| dc.description.abstract | Polyhydroxyalkanoate (PHA) synthase from a locally isolated Chromobacterium sp. USM2 (PhaCCs) exhibited superior polymerizing ability and broad in vivo substrate specificity with preferences for short chain length (SCL) [3- hydroxybutyrate (3HB) and 3-hydroxyvalerate (3HV)] and medium chain length (MCL) [3-hydroxyhexanoate (3HHx)] monomers. For further characterization of the synthase, a Strep2-tagged PhaCCs for expression in and purification from Escherichia coli, was constructed in this study. In vitro enzymatic assay revealed an activity of 253 ± 13 U/mg for polymerization of 3-hydroxybutyryl-coenzyme A (3HB-CoA), which was approximately fivefold higher than that of model PHAproducing strain Cupriavidus necator (39 ± 5 U/mg). Its activity for polymerization of 3-hydroxyvaleryl-coenzyme A was twice as great as that for 3HB-CoA, while corresponding activity for 3-hexanoyl-coenzyme A polymerization rivaled that of another SCL-MCL synthase, from Aeromonas caviae. This discovery prompted further characterization studies and application of the highly active synthase for in vivo PHA production. Here, the gene encoding the PHA synthase from Chromobacterium sp. USM2 (phaCCs) was used to replace the native PHA synthase gene in wild type C. necator by homologous recombination. The resultant strain showed improved productivity of flexible poly(3-hydroxybutyrate-co-3- hydroxyhexanoate) from crude palm kernel oil with concomitant good growth. Various approaches such as alteration of culture parameters, addition of precursor or inhibitor compounds and manipulation of biosynthetic pathway successfully increased the 3HHx monomer fraction, further enhancing flexibility of the copolymer. | en_US |
| dc.identifier.uri | http://hdl.handle.net/123456789/8318 | |
| dc.language.iso | en | en_US |
| dc.publisher | Universiti Sains Malaysia | en_US |
| dc.subject | Characterization of a highly active | en_US |
| dc.subject | Polyhydroxyalkanoate synthase | en_US |
| dc.title | Characterization Of A Highly Active Polyhydroxyalkanoate Synthase | en_US |
| dc.type | Thesis | en_US |
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