Poly(3-Hydroxybutyrate) (PHB) Polymerase PhaC1 and PHB Depolymerase PhaZa1 of Ralstonia eutropha Are Phosphorylated .

In this study, we screened poly(3-hydroxybutyrate) (PHB) synthase PhaC1 and PHB depolymerase PhaZa1 of for the presence of phosphorylated residues during the PHB accumulation and PHB degradation phases. Thr373 of PHB synthase PhaC1 was phosphorylated during the stationary growth phase but was not modified during the exponential and PHB accumulation phases. Ser35 of PHB depolymerase PhaZa1 was identified in the phosphorylated form during both the exponential and stationary growth phases.

Determination of Polyhydroxybutyrate (PHB) Content in Using Gas Chromatography and Nile Red Staining.

H16 produces and mobilizes (re-utilizes) intracellular polyhydroxybutyrate (PHB) granules during growth. This protocol describes the visualization of intracellular Nile red stained PHB granules and the quantification of PHB by gas chromatography. Our first method describes how to analyze PHB granules by fluorescence microscopy qualitatively.

Absence of ppGpp Leads to Increased Mobilization of Intermediately Accumulated Poly(3-Hydroxybutyrate) in Ralstonia eutropha H16.

In this study, we constructed a set of H16 strains with single, double, or triple deletions of the (p)ppGpp synthase/hydrolase (), (p)ppGpp synthase (), and/or polyhydroxybutyrate (PHB) depolymerase ( or ) gene, and we determined the impact on the levels of (p)ppGpp and on accumulated PHB. Mutants with deletions of both the and genes were unable to synthesize detectable amounts of (p)ppGpp and accumulated only minor amounts of PHB, due to PhaZa1-mediated depolymerization of PHB. In contrast, unusually high levels of PHB were found in strains in which the (p)ppGpp concentration was increased by the overexpression of (p)ppGpp synthase (SpoT2) and the absence of (p)ppGpp hydrolase.