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	Construction of pha-operon-defined knockout mutants of Pseudomonas putida KT2442 and their applications in poly(hydroxyalkanoate) production
	Ouyang, Shao-Ping ; Liu, Qian ; Fang, Lei ; Chen, Guo-Qiang
	2010-05-11 ; 2010-05-11
关键词	biopolyesters copolyesters PHB poly(hydroxyalkanoate)s Pseudomonas putida AEROMONAS-HYDROPHILA ENHANCED PRODUCTION POLYHYDROXYALKANOATE SYNTHASES ESCHERICHIA-COLI POLY(3-HYDROXYBUTYRATE-CO-3-HYDROXYHEXANOATE) CLONING GENES BIOSYNTHESIS COPOLYESTERS COPOLYMER Biochemistry & Molecular Biology Materials Science, Biomaterials Polymer Science
中文摘要	Pseudomonas putida KT2442 could accumulate medium-chain-length poly(hydroxyalkanoate)s (PHA) consisting of 3-hydroxyhexanoate, 3-hydroxyoctanoate, 3-hydroxydecanoate, and 3-hydroxydodecanoate from a wide range of carbon sources. In this study, the PHA synthase pha operon (phaC1-phaZ-phaC2) was knocked out and the vgb gene encoding vitreoscilla hemoglobin protein (VHb), which could enhance oxygen uptake rate especially at low oxygen concentration, was integrated into the P. putida KT2442 genome to replace the deleted fragment. The resulting mutant P. putida KTOY01 or gene-replaced mutant KTOY02 was used as the host to study PHA synthase properties and PHA production. Different PHA polymerase (PhaC) genes, phaC(Re) from Rastonia eutropha H16, phaC(Ac) from Aeromonas cavie, and phaC2(Ps) from Pseudomonas stutzeri 1317, were expressed in the mutant strains to test the PhaC enzyme substrate specificity. The result showed P. putida KTOY01 or KTOY02 could provide not only mcl PHA monomers but also 3-hydroxybutyrate from fatty acids, which may allow the production of copolyesters poly(3HB-co-mcl 3HA). Plasmid pCJY10 containing phaC2(Ps), phbA, and phbB genes encoding PHA polymerase, beta-ketothiolase, and acetoacetyl-CoA reductase, respectively, were transformed into P. putida KTOY01 and KTOY02. Shake-flask culture showed P. putida KTOY01 or KTOY02 (pCJY10) could accumulate poly(3HB-co-mcl 3HA) from glucose. The above result showed pha operon knockout mutant of P. putida KT2442 was,a very useful host of great potential not only for studying PhaC synthase, but also for microbial production of copolyesters poly(3HB-co-mcl 3HA), which is very difficult to obtain.
语种	英语 ; 英语
出版者	WILEY-V C H VERLAG GMBH ; WEINHEIM ; PO BOX 10 11 61, D-69451 WEINHEIM, GERMANY
内容类型	期刊论文
源URL	[http://hdl.handle.net/123456789/26507]
专题	清华大学
推荐引用方式 GB/T 7714	Ouyang, Shao-Ping,Liu, Qian,Fang, Lei,et al. Construction of pha-operon-defined knockout mutants of Pseudomonas putida KT2442 and their applications in poly(hydroxyalkanoate) production[J],2010, 2010.
APA	Ouyang, Shao-Ping,Liu, Qian,Fang, Lei,&Chen, Guo-Qiang.(2010).Construction of pha-operon-defined knockout mutants of Pseudomonas putida KT2442 and their applications in poly(hydroxyalkanoate) production..
MLA	Ouyang, Shao-Ping,et al."Construction of pha-operon-defined knockout mutants of Pseudomonas putida KT2442 and their applications in poly(hydroxyalkanoate) production".(2010).

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