Studies on poly-β-hydroxybutyrate synthase activity of Nostoc muscorum

Sharma, Lalita; Panda, Bhabatarini; Singh, Akhilesh Kumar; Mallick, Nirupama

doi:10.2323/jgam.52.209

Cited by 7 publications

(2 citation statements)

References 21 publications

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“…It has been reported that acetyl-P can substitute for phosphoenolpyruvate as the phospho donor for purified enzyme I of the phosphoenolpyru- VOL. 189, 2007 INTRACELLULAR ACETYL PHOSPHATE MEASUREMENT 5579 vate:carbohydrate phosphotransferase system (15) and can act as an allosteric effector for HPr kinase/phosphatase purified from the gram-positive organism Bacillus subtilis (41), for certain eukaryotic ATPases (7,22,49), and for poly-␤-hydroxybutyrate synthase from cyanobacteria (35,46). To the best of our knowledge, however, there have been no reports demonstrating that any of these interactions can occur in vivo.…”

Section: Discussionmentioning

confidence: 99%

The Intracellular Concentration of Acetyl Phosphate in Escherichia coli Is Sufficient for Direct Phosphorylation of Two-Component Response Regulators

et al. 2007

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Acetyl phosphate, the intermediate of the AckA-Pta pathway, acts as a global signal in Escherichia coli. Although acetyl phosphate clearly signals through two-component response regulators, it remains unclear whether acetyl phosphate acts as a direct phospho donor or functions through an indirect mechanism. We used two-dimensional thin-layer chromatography to measure the relative concentrations of acetyl phosphate, acetyl coenzyme A, ATP, and GTP over the course of the entire growth curve. We estimated that the intracellular concentration of acetyl phosphate in wild-type cells reaches at least 3 mM, a concentration sufficient to activate two-component response regulators via direct phosphoryl transfer.Bacterial cells must respond properly to diverse external and internal cues, a process that requires global signaling. The ideal global signal is metabolically inexpensive, short-lived, indicative only of significant changes, and capable of effecting the coordinated regulation of diverse cellular processes. Several lines of evidence suggest that the small molecule acetyl phosphate (acetyl-P) can serve as such a global signal.Acetyl-P is the high-energy, acid/base-labile intermediate of the reversible Pta-AckA pathway (Fig. 1). This pathway interconverts coenzyme A (HS-CoA), ATP, and acetate with acetyl coenzyme A (acetyl-CoA), ADP, and orthophosphate (P i ) (9, 43). The reversibility of this pathway permits both acetyl-CoA synthesis (acetate activation) and acetate evolution (acetogenesis). During acetogenesis, Pta synthesizes acetyl-P and HSCoA from acetyl-CoA and P i , while AckA generates ATP from acetyl-P and ADP. Simultaneously, AckA produces acetate, which cells excrete into the environment. Thus, the steadystate concentration of acetyl-P depends upon the rate of its formation catalyzed by Pta and the rate of its degradation catalyzed by AckA (for reviews, see references 45 and 59).Acetyl-P has been proposed to act as a global regulator by direct phosphorylation of response regulators (RRs) of the family of two-component signal transduction (2CST) pathways (34, 55). The simplest of these 2CST pathways consists of a histidine kinase (HK) and an RR. Using ATP as its phosphoryl donor, the HK autophosphorylates a conserved histidine residue. In turn, the RR autophosphorylates a conserved aspartate residue, using its phosphorylated cognate HK as its phosphoryl donor. Experiments performed in vitro have clearly demonstrated that acetyl-P can donate its phosphoryl group to purified RRs but not to HKs (for reviews, see references 52 and 57). This ability arises from its capacity to store energy. Acetyl-P possesses a larger ⌬G°of hydrolysis (Ϫ43.3 kJ/mol) than ATP possesses (Ϫ30.5 kJ/mol in complex with Mg 2ϩ ). This difference forms the basis for the role of acetyl-P in generating ATP (for a review, see reference 59).Although acetyl-P can function as a phosphoryl donor in vitro, its ability to function in vivo as a global signal has remained essentially unproven, despite a wealth of seemingly supportive data (for a r...

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Section: Discussionmentioning

confidence: 99%

The Intracellular Concentration of Acetyl Phosphate in Escherichia coli Is Sufficient for Direct Phosphorylation of Two-Component Response Regulators

et al. 2007

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“…The activity of PHB synthase was determined using the method of Sharma et al (2006) who measured the absorbance of CoA at A 412nm . The activity of glucose-6-phosphate dehydrogenase (G6PD) was analysed by measuring the released NADPH at A 340nm (Sagisaka 1972).…”

Section: Measurement Of Enzyme Activitiesmentioning

confidence: 99%

Effect of ethanol and hydrogen peroxide on poly(3-hydroxybutyrate) biosynthetic pathway in Cupriavidus necator H16

Obruča

Márová

et al. 2010

World J Microbiol Biotechnol

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Exposition of Cupriavidus necator to ethanol or hydrogen peroxide at the beginning of the stationary phase increases poly(3-hydroxybutyrate) (PHB) yields about 30%. Hydrogen peroxide enhances activity of pentose phosphate pathway that probably consequently increases intracellular ratio NADPH/NADP(+). This effect leads to stimulation of the flux of acetyl-CoA into PHB biosynthetic pathway and to an increase of enzymatic activities of β-ketothiolase and acetoacetyl-CoA reductase while activity of PHB synthase remains uninfluenced. During ethanol metabolisation, in which alcohol dehydrogenase is involved, acetyl-CoA and reduced coenzymes NAD(P)H are formed. These metabolites could again slightly inhibit TCA cycle while flux of acetyl-CoA into PHB biosynthetic pathway is likely to be supported. As a consequence of TCA cycle inhibition also less free CoA is formed. Similarly with hydrogen peroxide, activities of β-ketothiolase and acetoacetyl-CoA reductase are increased which results in over-production of PHB. Molecular weight of PHB produced under stress conditions was significantly higher as compared to control cultivation. Particular molecular weight values were dependent on stress factor concentrations. This could indicate some interconnection among activities of β-ketothiolase, acetoacetyl-CoA reductase and PHB molecular weight control in vivo.

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