Accumulation of the PhaP Phasin of
            <i>Ralstonia eutropha</i>
            Is Dependent on Production of Polyhydroxybutyrate in Cells

York, Gregory M.; Junker, Björn H.; Stubbe, JoAnne; Sinskey, Anthony J.

doi:10.1128/jb.183.14.4217-4226.2001

Cited by 109 publications

(124 citation statements)

References 35 publications

Supporting

Mentioning

113

Contrasting

Unclassified

Order By: Relevance

“…PHA-forming cells synthesize phasins only in amounts that can be bound to the granules (52). Therefore, the occurrence of phasins can be used as a marker for PHA synthesis (55). In the present proteomic analysis, the levels of PhbB and PhaC were increased 3.0-fold during growth with 0.74 mM toluene.…”

Section: Resultsmentioning

confidence: 61%

Solvent Stress Response of the Denitrifying Bacterium “ Aromatoleum aromaticum ” Strain EbN1

Trautwein

Kühner

Wöhlbrand

et al. 2008

Appl Environ Microbiol

View full text Add to dashboard Cite

The denitrifying betaproteobacterium "Aromatoleum aromaticum" strain EbN1 degrades several aromatic compounds, including ethylbenzene, toluene, p-cresol, and phenol, under anoxic conditions. The hydrophobicity of these aromatic solvents determines their toxic properties. Here, we investigated the response of strain EbN1 to aromatic substrates at semi-inhibitory (about 50% growth inhibition) concentrations under two different conditions: first, during anaerobic growth with ethylbenzene (0.32 mM) or toluene (0.74 mM); and second, when anaerobic succinate-utilizing cultures were shocked with ethylbenzene (0.5 mM), toluene (1.2 mM), p-cresol (3.0 mM), and phenol (6.5 mM) as single stressors or as a mixture (total solvent concentration, 2.7 mM). Under all tested conditions impaired growth was paralleled by decelerated nitrate-nitrite consumption. Additionally, alkylbenzene-utilizing cultures accumulated poly(3-hydroxybutyrate) (PHB) up to 10% of the cell dry weight. These physiological responses were also reflected on the proteomic level (as determined by two-dimensional difference gel electrophoresis), e.g., up-regulation of PHB granule-associated phasins, cytochrome cd 1 nitrite reductase of denitrification, and several proteins involved in oxidative (e.g., SodB) and general (e.g., ClpB) stress responses.

show abstract

Section: Resultsmentioning

confidence: 61%

Solvent Stress Response of the Denitrifying Bacterium “ Aromatoleum aromaticum ” Strain EbN1

Trautwein

Kühner

Wöhlbrand

et al. 2008

Appl Environ Microbiol

View full text Add to dashboard Cite

show abstract

“…It may be assumed that PhaP1 possesses similar regions at least in the mature protein. In contrast to oleosins of plant seeds, PhaP1 is directly involved in the regulation of the size, quantity, and formation of PHA inclusions and it was shown that the amount of PhaP1 parellels the amount of PHA in the cells (108,126,143,148). The presence or absence of this protein interferes with the shape and number of PHA inclusions, as revealed in mutants overexpressing or lacking PhaP1 (143).…”

Section: Proteins Associated With Prokaryotic Lipid Inclusionsmentioning

confidence: 99%

Neutral Lipid Bodies in Prokaryotes: Recent Insights into Structure, Formation, and Relationship to Eukaryotic Lipid Depots

2005

View full text Add to dashboard Cite

Many prokaryotes are able to accumulate large amounts of lipophilic compounds as inclusion bodies in the cytoplasm. Members of most genera synthesize polymeric lipids such as poly(3-hydroxybutyrate) (PHB) or other polyhydroxyalkanoates (PHAs) (125), whereas accumulation of triacylglycerols (TAGs) and wax esters (WEs) in intracellular lipid-bodies is a property of only a few prokaryotes (10). Like the formation of PHAs, TAG and WE biosynthesis is also promoted in response to stress imposed on the cells and during imbalanced growth, for example by nitrogen limitation, if an abundant carbon source is present at the same time. All these lipids act as storage compounds for energy and carbon needed for maintenance of metabolism and synthesis of cellular metabolites during starvation and in particular if growth resumes. Although neutral lipid metabolism in bacteria, especially WE biosynthesis, has attracted increasing biotechnological interest, there has so far been little interest in medical research in the formation of prokaryotic lipid bodies. However, recent findings suggest that lipid body formation and accumulation of TAGs play an important role in the metabolism of the pathogenic bacteria like Mycobacterium tuberculosis (37,52).In contrast to the restricted occurrence of storage TAGs in prokaryotes, intracellular TAGs are widespread in many eukaryotes (34,35,40,53,65,106,112,132,133). In eukaryotes, storage lipids are also deposited as lipid bodies. Their structure and formation have recently been reviewed in detail by several authors, but additional information on prokaryotic lipid-bodies has been only superficial (98,100,151). In contrast, intracellular accumulation of WEs as a storage lipid is a great exception in eukaryotes and occurs only in jojoba (Simmondsia chinensis) (147), and PHAs are not at all synthesized as a storage compound in eukaryotes.The number of studies investigating the enzymatic and structural fundamentals of storage lipid metabolism in bacteria has increased drastically in the last years, providing the knowledge that storage lipids in bacteria have a similar important metabolic function like in eukaryotes. This review will focus on prokaryotic neutral lipid bodies in comparison with the currently discussed models for lipid body structure and their formation in eukaryotes and with PHA inclusions in prokaryotes.

show abstract

“…Standard approaches were used for DNA preparation and manipulation (1). Genomic DNA was extracted from R. eutropha with modifications previously described (23). PCR products were routinely cloned into pCR2.1-TOPO using a TOPO TA cloning kit (Invitrogen Corp., Carlsbad, Calif.).…”

Section: Methodsmentioning

confidence: 99%

Ralstonia eutrophaH16 Encodes Two and Possibly Three Intracellular Poly[d-(−)-3-Hydroxybutyrate] Depolymerase Genes

York¹,

Lupberger²,

et al. 2003

Self Cite

View full text Add to dashboard Cite

Intracellular poly[D-(؊)-3-hydroxybutyrate] (PHB) depolymerases degrade PHB granules to oligomers and monomers of 3-hydroxybutyric acid.Recently an intracellular PHB depolymerase gene (phaZ1) from Ralstonia eutropha was identified. We now report identification of candidate PHB depolymerase genes from R. eutropha, namely, phaZ2 and phaZ3, and their characterization in vivo. phaZ1 was used to identify two candidate depolymerase genes in the genome of Ralstonia metallidurans. phaZ1 and these genes were then used to design degenerate primers. These primers and PCR methods on the R. eutropha genome were used to identify two new candidate depolymerase genes in R. eutropha: phaZ2 and phaZ3. Inverse PCR methods were used to obtain the complete sequence of phaZ3, and library screening was used to obtain the complete sequence of phaZ2. PhaZ1, PhaZ2, and PhaZ3 share ϳ30% sequence identity. The function of PhaZ2 and PhaZ3 was examined by generating R. eutropha H16 deletion strains (⌬phaZ1, ⌬phaZ2, ⌬phaZ3, ⌬phaZ1⌬phaZ2, ⌬phaZ1⌬phaZ3, ⌬phaZ2⌬phaZ3, and ⌬phaZ1⌬phaZ2⌬phaZ3). These strains were analyzed for PHB production and utilization under two sets of conditions. When cells were grown in rich medium, PhaZ1 was sufficient to account for intracellular PHB degradation. When cells that had accumulated ϳ80% (cell dry weight) PHB were subjected to PHB utilization conditions, PhaZ1 and PhaZ2 were sufficient to account for PHB degradation. PhaZ2 is thus suggested to be an intracellular depolymerase. The role of PhaZ3 remains to be established.Polyhydroxyalkanoates (PHAs) are polyoxoesters produced by a wide range of bacteria when they find themselves in an environment with an available carbon source but limited in additional nutrient(s) required for growth (9). The shortchain-length PHAs, where R is a methyl or ethyl, have properties of thermoplastics and are biodegradable (Fig. 1). Much effort has focused on understanding the biology of PHA homeostasis for several reasons. First, this understanding could lead to expression of the appropriate gene set in heterologous systems to make PHA production economically competitive with oil-based polymers. Second, understanding PHA homeostasis serves as a paradigm for understanding the mechanism of homopolymerization reactions in which the product undergoes a phase transition during its formation, generating insoluble inclusions (granules). The intracellular PHAs can be degraded when the bacteria require carbon but are in otherwise nutrientreplete conditions, and the monomers and energy released can be reused to allow the bacteria to grow (17). The insoluble PHA granules must, therefore, be biosynthesized in a controlled fashion to facilitate enzymatic degradation. A variety of proteins associated with PHA homeostasis have been identified and are being characterized (3,8,10,12,20,22). As part of our research to understand the mechanisms that control polymer size and reuse, we have been interested in identifying the intracellular depolymerases that degrade poly[D-(Ϫ)-3-hydroxybutyrate] (PHB) w...

show abstract

Accumulation of the PhaP Phasin of Ralstonia eutropha Is Dependent on Production of Polyhydroxybutyrate in Cells

Cited by 109 publications

References 35 publications

Solvent Stress Response of the Denitrifying Bacterium “ Aromatoleum aromaticum ” Strain EbN1

Solvent Stress Response of the Denitrifying Bacterium “ Aromatoleum aromaticum ” Strain EbN1

Neutral Lipid Bodies in Prokaryotes: Recent Insights into Structure, Formation, and Relationship to Eukaryotic Lipid Depots

Ralstonia eutrophaH16 Encodes Two and Possibly Three Intracellular Poly[d-(−)-3-Hydroxybutyrate] Depolymerase Genes

Contact Info

Product

Resources

About