Analyses of binding sequences of the PhaR protein of Rhodobacter sphaeroides FJ1

Chou, Min-En; Yang, Ming‐Der

doi:10.1111/j.1574-6968.2009.01836.x

Cited by 10 publications

(9 citation statements)

References 25 publications

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“…A similar sequence (CTGC[N] 3 GCAG) was also described in R. sphaeroides FJ1 as the DNA-binding site for the regulator PhaR [41]. Both sequences show two highly conserved triplets (TGC and GCA) which seem to be essential for DNA-binding of R. sphaeroides PhaR [41]. …”

Section: Resultsmentioning

confidence: 89%

Identification and characterization of PhbF: A DNA binding protein with regulatory role in the PHB metabolism of Herbaspirillum seropedicae SmR1

et al. 2011

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BackgroundHerbaspirillum seropedicae SmR1 is a nitrogen fixing endophyte associated with important agricultural crops. It produces polyhydroxybutyrate (PHB) which is stored intracellularly as granules. However, PHB metabolism and regulatory control is not yet well studied in this organism.ResultsIn this work we describe the characterization of the PhbF protein from H. seropedicae SmR1 which was purified and characterized after expression in E. coli. The purified PhbF protein was able to bind to eleven putative promoters of genes involved in PHB metabolism in H. seropedicae SmR1. In silico analyses indicated a probable DNA-binding sequence which was shown to be protected in DNA footprinting assays using purified PhbF. Analyses using lacZ fusions showed that PhbF can act as a repressor protein controlling the expression of PHB metabolism-related genes.ConclusionsOur results indicate that H. seropedicae SmR1 PhbF regulates expression of phb-related genes by acting as a transcriptional repressor. The knowledge of the PHB metabolism of this plant-associated bacterium may contribute to the understanding of the plant-colonizing process and the organism's resistance and survival in planta.

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Section: Resultsmentioning

confidence: 89%

Identification and characterization of PhbF: A DNA binding protein with regulatory role in the PHB metabolism of Herbaspirillum seropedicae SmR1

et al. 2011

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“…Among various carbon sources, acetate yielded the highest PHB production [17]. Moreover, PHB productivity can be greatly improved within R. sphaeroides, since it is possible to enhance the performance of strains using well-established genetic engineering tools and by integrating advances in previous studies [5,[18][19][20].…”

Section: Introductionmentioning

confidence: 99%

Molecular Profiling and Optimization Studies for Growth and PHB Production Conditions in Rhodobacter sphaeroides

et al. 2020

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In the recent climate change regime, industrial demand for renewable materials to replace petroleum-derived polymers continues to rise. Of particular interest is polyhydroxybutyrate (PHB) as a substitute for polypropylene. Accumulating evidence indicates that PHB is highly produced as a carbon storage material in various microorganisms. The effects of growth conditions on PHB production have been widely studied in chemolithotrophs, particularly in Rhodobacter. However, the results on PHB production in Rhodobacter have been somewhat inconsistent due to different strains and experimental conditions, and it is currently unclear how diverse environmental factors are linked with PHB production. Here, we report optimized growth conditions for PHB production and show that the growth conditions are closely related to reactive oxygen species (ROS) regulation. PHB accumulates in cells up to approximately 50% at the highest level under dark-aerobic conditions as opposed to light aerobic/anaerobic conditions. According to the time-course, PHB contents increased at 48 h and then gradually decreased. When observing the effect of temperature and medium composition on PHB production, 30 °C and a carbon/nitrogen ratio of 9:1 or more were found to be most effective. Among PHB biosynthetic genes, PhaA and PhaB are highly correlated with PHB production, whereas PhaC and PhaZ showed little change in overall expression levels. We found that, while the amount of hydrogen peroxide in cells under dark conditions was relatively low compared to the light conditions, peroxidase activities and expression levels of antioxidant-related genes were high. These observations suggest optimal culture conditions for growth and PHB production and the importance of ROS-scavenging signaling with regard to PHB production.

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“…However, despite the attractive characteristics of PNS bacteria as a host microorganism for bioproduction, only a few studies about PHA including poly (3-hydroxybutyrate) (PHB) biosynthesis using genetic engineering techniques have been reported so far [15–18]. Furthermore, these studies did not focus on improving PHA production, but on describing the genetic function and PHA biosynthesis in PNS bacteria.…”

Section: Introductionmentioning

confidence: 99%

Disruption of poly (3-hydroxyalkanoate) depolymerase gene and overexpression of three poly (3-hydroxybutyrate) biosynthetic genes improve poly (3-hydroxybutyrate) production from nitrogen rich medium by Rhodobacter sphaeroides

Kobayashi

2019

Microb Cell Fact

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Background Due to various environmental problems, biodegradable polymers such as poly (3-hydroxybutyrate) (PHB) have gained much attention in recent years. Purple non-sulfur (PNS) bacteria have various attractive characteristics useful for environmentally harmless PHB production. However, production of PHB by PNS bacteria using genetic engineering has never been reported. This study is the first report of a genetically engineered PNS bacterial strain with a high PHB production. Results We constructed a poly (3-hydroxyalkanoate) depolymerase ( phaZ ) gene-disrupted Rhodobacter sphaeroides HJ strain. This R. sphaeroides HJΔ phaZ (pLP-1.2) strain showed about 2.9-fold higher volumetric PHB production than that of the parent HJ (pLP-1.2) strain after 5 days of culture. The HJΔ phaZ strain was further improved for PHB production by constructing strains overexpressing each of the eight genes including those newly found and annotated as PHB biosynthesis genes in the KEGG GENES Database. Among these constructed strains, all of gene products exhibited annotated enzyme activities in the recombinant strain cells, and HJΔ phaZ ( phaA3 ), HJΔ phaZ ( phaB2 ), and HJΔ phaZ ( phaC1 ) showed about 1.1-, 1.1-, and 1.2-fold higher volumetric PHB production than that of the parent HJΔ phaZ (pLP-1.2) strain. Furthermore, we constructed a strain that simultaneously overexpresses all three phaA3 , phaB2 , and phaC1 genes; this HJΔ phaZ ( phaA3 / phaB2 / phaC1 ) strain showed about 1.7- to 3.9-fold higher volumetric PHB production (without ammonium sulfate; 1.88 ± 0.08 g l −1 and with 100 mM ammonium sulfate; 0.99 ± 0.05 g l −1 ) than those of the parent HJ (pLP-1.2) strain grown under nitrogen limited and rich conditions, respectively. Conclusion In this study, we identified eight different genes involved in PHB biosynthesis in the genome of R. sphaeroides 2.4.1, and revealed that their overexpression increased PHB accumulation in an R. sphaeroides HJ strain. In addition, we demonstrated the effectiveness of a phaZ disruption for high PHB accumulation, especially under nitrogen rich conditions. Furthermore, we showed that PNS bacteria may have some unidentified genes involved in poly (3-hydroxyalkanoates) (PHA) biosynthesis. Our findings could lead to further improvement of environmentally harmless PHA production techniques ...

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Analyses of binding sequences of the PhaR protein of Rhodobacter sphaeroides FJ1

Cited by 10 publications

References 25 publications

Identification and characterization of PhbF: A DNA binding protein with regulatory role in the PHB metabolism of Herbaspirillum seropedicae SmR1

Identification and characterization of PhbF: A DNA binding protein with regulatory role in the PHB metabolism of Herbaspirillum seropedicae SmR1

Molecular Profiling and Optimization Studies for Growth and PHB Production Conditions in Rhodobacter sphaeroides

Disruption of poly (3-hydroxyalkanoate) depolymerase gene and overexpression of three poly (3-hydroxybutyrate) biosynthetic genes improve poly (3-hydroxybutyrate) production from nitrogen rich medium by Rhodobacter sphaeroides

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