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In vitro synthesis of polyhydroxyalkanoate (PHA) incorporating lactate (LA) with a block sequence by using a newly engineered thermostable PHA synthase from Pseudomonas sp. SG4502 with acquired LA-polymerizing activity
| Title: | In vitro synthesis of polyhydroxyalkanoate (PHA) incorporating lactate (LA) with a block sequence by using a newly engineered thermostable PHA synthase from Pseudomonas sp. SG4502 with acquired LA-polymerizing activity |
| Authors: | Tajima, Kenji Browse this author →KAKEN DB | | Han, Xuerong Browse this author | | Satoh, Yasuharu Browse this author →KAKEN DB | | Ishii, Ayako Browse this author | | Araki, Yuji Browse this author | | Munekata, Masanobu Browse this author →KAKEN DB | | Taguchi, Seiichi Browse this author →KAKEN DB |
| Keywords: | Polyhydroxyalkanoate (PHA) | | Thermal stability | | Block copolymer | | LA-polymerizing enzyme (LPE) | | In vitro polymerization | | Pseudomonas sp. SG4502 |
| Issue Date: | Apr-2012 |
| Publisher: | Springer Berlin / Heidelberg |
| Journal Title: | Applied Microbiology and Biotechnology |
| Volume: | 94 |
| Issue: | 2 |
| Start Page: | 365 |
| End Page: | 376 |
| Publisher DOI: | 10.1007/s00253-011-3840-z |
| PMID: | 22249718 |
| Abstract: | Recently, we succeeded in isolating a thermotolerant bacterium, Pseudomonas sp. SG4502, which is capable of accumulating polyhydroxyalkanoate (PHA) even at 55℃, as a source of thermostable enzymes. In this study, we cloned a pha locus from the bacterium and identified two genes encoding PHA synthases (PhaC1_[SG] and PhaC2_[SG]). Two mutations, Ser324Thr and Gln480Lys, corresponding to those of a lactate (LA)-polymerizing enzyme (LPE) from mesophilic Pseudomonas sp. 61-3 were introduced into PhaC1_[SG] to evaluate the potential of the resulting protein as a "thermostable LPE". The mutated PhaC1_[SG] [PhaC1_[SG](STQK)] showed high thermal stability in synthesizing P(LA-co-3HB) in an in vitro reaction system under a range of high temperatures. Requirement of 3HBCoA as a priming unit for LA polymerization by the LPE has been suggested in both of the in vitro and in vivo experiments. Based on the finding, the PhaC1_[SG](STQK)-mediated synthesis of a LA-based copolymer with a block sequence was achieved in the in vitro system by sequential feeding of the corresponding two substrates. This in vitro reaction system using the thermostable LPE provides us with a versatile way to synthesize the various types of LA-based copolymers with desired sequence patterns, random or block, depending on the way of supplying hydroxyalkanoates (mixed or sequential feeding). |
| Rights: | The original publication is available at www.springerlink.com |
| Type: | article (author version) |
| URI: | http://hdl.handle.net/2115/52295 |
| Appears in Collections: | 工学院・工学研究院 (Graduate School of Engineering / Faculty of Engineering) > 雑誌発表論文等 (Peer-reviewed Journal Articles, etc)
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Submitter: 田島 健次
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