Enhanced production of medium-chain-length polyhydroxyalkanoates (PHA) by PHA depolymerase knockout mutant of Pseudomonas putida KT2442

Cai, Lei; Yuan, Meiqing; Feng, Lian‐Shun; Jia, Jian; Chen, Guoqiang

doi:10.1016/j.biortech.2008.11.020

Cited by 107 publications

(49 citation statements)

References 35 publications

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“…However, the polymers differed in length: the molecular weights (Mw and Mn values) of the polymers from the PpU parental strain and the PpU 10–33 (PhaC2 polymerase hyperexpressing construct) were similar, ranging from 126–142 and 74–77 kDa respectively, whereas those from the PhaZ knockout were considerably lower, 96 and 50 kDa respectively. Although this result may seem to be counter‐intuitive, it is consistent with observations of Cai and colleagues () and Solaiman and colleagues (), who also found lower polymer lengths in phaZ mutants. Thus, we cannot rule out the possibility that the depolymerase also plays a direct or indirect role in chain length determination and/or in PHA synthesis itself.…”

Section: Resultsmentioning

confidence: 99%

Tight coupling of polymerization and depolymerization of polyhydroxyalkanoates ensures efficient management of carbon resources in Pseudomonas putida

Arias

Bassas-Galià

Molinari

et al. 2013

Microbial Biotechnology

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SummaryEnvironmental microbes oscillate between feast and famine and need to carefully manage utilization, storage and conversion of reserve products to exploitable sources of carbon and energy. Polyhydroxyalkanoates (PHAs) are storage polymers that serve bacteria as sources of food materials under physiological conditions of carbon demand. In order to obtain insights into the role of PHA depolymerase (PhaZ) and its relationship to a PHA polymerase (PhaC2) in the carbon management activity of Pseudomonas putida strain U, we created a polymerase hyperexpression strain and a depolymerase knockout mutant of this strain, and examined their synthesis of PHA and expression of their PHA genes. This study revealed that hyperexpression of PhaC2 led to the accumulation of higher amounts of PHA (44%wt) than in the wild-type strain (24%wt) after 24 h of cultivation, which then returned to wild-type levels by 48 h, as a result of elevated depolymerization. The phaZ mutant, however, accumulated higher levels of PHA than the parental strain (62%wt), which were maintained for at least 96 h. Transcriptional analysis of the pha cluster by RT-PCR revealed that PHA operon proteins, including depolymerase, are expressed from the beginning of the growth phase. Hyperexpression of the PhaC2 polymerase was accompanied by an increase in the expression of the PhaZ depolymerase and a decrease in expression of another PHA polymerase, PhaC1. This suggests tight regulatory coupling of PHA polymerase and depolymerase activities that act in synergy, and in concert with other PHA proteins, to provide dynamic PHA granule synthesis and remodelling that rapidly and sensitively respond to changes in availability of carbon and the physiological-metabolic needs of the cell, to ensure optimal carbon resource management.

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Section: Resultsmentioning

confidence: 99%

Tight coupling of polymerization and depolymerization of polyhydroxyalkanoates ensures efficient management of carbon resources in Pseudomonas putida

Arias

Bassas-Galià

Molinari

et al. 2013

Microbial Biotechnology

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“…However, for industrial production of PHA, degradation is not favored, because it leads to the waste of carbon sources and consequently reduces PHA accumulation [47,48]. For example, it has been reported that the knockout of phaZ in P. putida KT2442 inhibited the decrease of PHA content during batch fermentation processes [47]. For Halomonas sp.…”

Section: Discussionmentioning

confidence: 99%

Comparative genomics study of polyhydroxyalkanoates (PHA) and ectoine relevant genes from Halomonas sp. TD01 revealed extensive horizontal gene transfer events and co-evolutionary relationships

et al. 2011

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BackgroundHalophilic bacteria have shown their significance in industrial production of polyhydroxyalkanoates (PHA) and are gaining more attention for genetic engineering modification. Yet, little information on the genomics and PHA related genes from halophilic bacteria have been disclosed so far.ResultsThe draft genome of moderately halophilic bacterium, Halomonas sp. TD01, a strain of great potential for industrial production of short-chain-length polyhydroxyalkanoates (PHA), was analyzed through computational methods to reveal the osmoregulation mechanism and the evolutionary relationship of the enzymes relevant to PHA and ectoine syntheses. Genes involved in the metabolism of PHA and osmolytes were annotated and studied in silico. Although PHA synthase, depolymerase, regulator/repressor and phasin were all involved in PHA metabolic pathways, they demonstrated different horizontal gene transfer (HGT) events between the genomes of different strains. In contrast, co-occurrence of ectoine genes in the same genome was more frequently observed, and ectoine genes were more likely under coincidental horizontal gene transfer than PHA related genes. In addition, the adjacent organization of the homologues of PHA synthase phaC1 and PHA granule binding protein phaP was conserved in the strain TD01, which was also observed in some halophiles and non-halophiles exclusively from γ-proteobacteria. In contrast to haloarchaea, the proteome of Halomonas sp. TD01 did not show obvious inclination towards acidity relative to non-halophilic Escherichia coli MG1655, which signified that Halomonas sp. TD01 preferred the accumulation of organic osmolytes to ions in order to balance the intracellular osmotic pressure with the environment.ConclusionsThe accessibility of genome information would facilitate research on the genetic engineering of halophilic bacteria including Halomonas sp. TD01.

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“…2A). Similarly, the inactivation of phaZ in P. putida KT2442 (Cai et al, 2009) and P. fluorescens BM07 (Choi et al, 2010) strains led to an increased accumulation of PHA. As expected, PAO1 phaZ mutant grown on fructose (Fig.…”

Section: Strainmentioning

confidence: 94%

Metabolic relationship between polyhydroxyalkanoic acid and rhamnolipid synthesis in Pseudomonas aeruginosa: Comparative 13C NMR analysis of the products in wild-type and mutants

Choi

García-Gutiérrez

et al. 2011

Journal of Biotechnology

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Enhanced production of medium-chain-length polyhydroxyalkanoates (PHA) by PHA depolymerase knockout mutant of Pseudomonas putida KT2442

Cited by 107 publications

References 35 publications

Tight coupling of polymerization and depolymerization of polyhydroxyalkanoates ensures efficient management of carbon resources in Pseudomonas putida

Tight coupling of polymerization and depolymerization of polyhydroxyalkanoates ensures efficient management of carbon resources in Pseudomonas putida

Comparative genomics study of polyhydroxyalkanoates (PHA) and ectoine relevant genes from Halomonas sp. TD01 revealed extensive horizontal gene transfer events and co-evolutionary relationships

Metabolic relationship between polyhydroxyalkanoic acid and rhamnolipid synthesis in Pseudomonas aeruginosa: Comparative 13C NMR analysis of the products in wild-type and mutants

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