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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

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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)

Submitter: 田島 健次

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