Smart polyhydroxyalkanoate nanobeads by protein based functionalization

Dinjaski, Nina; Prieto, María Auxiliadora

doi:10.1016/j.nano.2015.01.018

Cited by 53 publications

(39 citation statements)

References 137 publications

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“…The properties and functionalities of the PHAs depend on their monomer composition: whereas scl‐PHAs show thermoplastic properties similar to polypropylene, mcl‐PHAs display elastic features similar to rubber or elastomer (Keshavarz and Roy, ; Park et al ., ). Applications of PHAs in the industry are widespread, ranging from the manufacturing of packages and covers to the generation of enantiomeric pure chemicals (Philip et al ., ) or as protein immobilization supports (Draper and Rehm, ; Dinjaski and Prieto, ; Hay et al ., ). Of significant relevance is the implementation of PHAs in the biomedical discipline, especially supported by the recent FDA approval for P(4HB) to be used as suture material (Tepha Inc., MA, USA).…”

Section: Introductionmentioning

confidence: 99%

Polyhydroxyalkanoate‐associated phasins as phylogenetically heterogeneous, multipurpose proteins

Maestro

Sanz

2017

Microbial Biotechnology

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SummaryPolyhydroxyalkanoates (PHAs) are natural polyesters of increasing biotechnological importance that are synthesized by many prokaryotic organisms as carbon and energy storage compounds in limiting growth conditions. PHAs accumulate intracellularly in form of inclusion bodies that are covered with a proteinaceous surface layer (granule‐associated proteins or GAPs) conforming a network‐like surface of structural, metabolic and regulatory polypeptides, and configuring the PHA granules as complex and well‐organized subcellular structures that have been designated as ‘carbonosomes’. GAPs include several enzymes related to PHA metabolism (synthases, depolymerases and hydroxylases) together with the so‐called phasins, an heterogeneous group of small‐size proteins that cover most of the PHA granule and that are devoid of catalytic functions but nevertheless play an essential role in granule structure and PHA metabolism. Structurally, phasins are amphiphilic proteins that shield the hydrophobic polymer from the cytoplasm. Here, we summarize the characteristics of the different phasins identified so far from PHA producer organisms and highlight the diverse opportunities that they offer in the Biotechnology field.

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Section: Introductionmentioning

confidence: 99%

Polyhydroxyalkanoate‐associated phasins as phylogenetically heterogeneous, multipurpose proteins

Maestro

Sanz

2017

Microbial Biotechnology

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“…Two of its properties make R. eutropha famous: on the one hand, R. eutropha is able to grow chemolithoautotrophically with hydrogen and carbon dioxide as the electron donor and acceptor, respectively (a Knallgas bacterium), and on the other hand, it accumulates large amounts (Ͼ80% [dry weight] of the cell) of PHB. PHA produced by R. eutropha or related species are commercially available biopolymers and can be used for different purposes (10,11).…”

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confidence: 99%

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

Juengert

Borisova

Mayer

et al. 2017

Appl Environ Microbiol

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In this study, we constructed a set of Ralstonia eutropha H16 strains with single, double, or triple deletions of the (p)ppGpp synthase/hydrolase (spoT1), (p)ppGpp synthase (spoT2), and/or polyhydroxybutyrate (PHB) depolymerase (phaZa1 or phaZa3) gene, and we determined the impact on the levels of (p)ppGpp and on accumulated PHB. Mutants with deletions of both the spoT1 and spoT2 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. Determination of (p)ppGpp levels in wild-type R. eutropha under different growth conditions and induction of the stringent response by amino acid analogs showed that the concentrations of (p)ppGpp during the growth phase determine the amount of PHB remaining in later growth phases by influencing the efficiency of the PHB mobilization system in stationary growth. The data reported for a previously constructed ΔspoT2 strain (C. J. Brigham, D. R. Speth, C. Rha, and A. J. Sinskey, Appl Environ Microbiol 78:8033-8044, 2012, https://doi.org/ 10.1128/AEM.01693-12) were identified as due to an experimental error in strain construction, and our results are in contrast to the previous indication that the spoT2 gene product is essential for PHB accumulation in R. eutropha. IMPORTANCE Polyhydroxybutyrate (PHB) is an important intracellular carbon and energy storage compound in many prokaryotes and helps cells survive periods of starvation and other stress conditions. Research activities in several laboratories over the past 3 decades have shown that both PHB synthase and PHB depolymerase are constitutively expressed in most PHB-accumulating bacteria, such as Ralstonia eutropha. This implies that PHB synthase and depolymerase activities must be well regulated in order to avoid a futile cycle of simultaneous PHB synthesis and PHB degradation (mobilization). Previous reports suggested that the stringent response in Rhizobium etli and R. eutropha is involved in the regulation of PHB metabolism. However, the levels of (p)ppGpp and the influence of those levels on PHB accumulation and PHB mobilization have not yet been determined for any PHB-accumulating species. In this study, we optimized a (p)ppGpp extraction procedure and a highperformance liquid chromatography-mass spectrometry (HPLC-MS)-based detection method for the quantification of (p)ppGpp in R. eutropha. This enabled us to study the relationship between the concentrations of (p)ppGpp and the accumulated levels of PHB in the wild type and in several constructed mutant strains. We show that overproduction of the alarmone (p)ppGpp correlated with reduced growth and mas-

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“…While it is generally accepted that several proteins are part of the PHB/PHA granule surface layer in vivo , the presence or absence of phospholipids on the PHB/PHA granule surface is controversially discussed45850555657. The basis for the assumption that phospholipids are present in the surface layer of PHB granules is the identification of phosphatidic acid and at least one other lipid-like (acetone extractable) compound in purified PHB granules of Bacillus megaterium almost 50 years ago55 and the detection of phosphatidyl-ethanolamine, phosphatidyl-glycerol, diphosphatidyl-glycerol and a fourth not-identified compound (possibly phosphatidyl-serine) in isolated native PHB granules of R. eutropha (at that time designated as Alcaligenes eutrophus )58.…”

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confidence: 99%

Polyhydroxyalkanoate (PHA) Granules Have no Phospholipids

Bresan¹,

Sznajder²,

Hauf

et al. 2016

Sci Rep

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Polyhydroxybutyrate (PHB) granules, also designated as carbonosomes, are supra-molecular complexes in prokaryotes consisting of a PHB polymer core and a surface layer of structural and functional proteins. The presence of suspected phospholipids in the surface layer is based on in vitro data of isolated PHB granules and is often shown in cartoons of the PHB granule structure in reviews on PHB metabolism. However, the in vivo presence of a phospholipid layer has never been demonstrated. We addressed this topic by the expression of fusion proteins of DsRed2EC and other fluorescent proteins with the phospholipid-binding domain (LactC2) of lactadherin in three model organisms. The fusion proteins specifically localized at the cell membrane of Ralstonia eutropha but did not co-localize with PHB granules. The same result was obtained for Pseudomonas putida, a species that accumulates another type of polyhydroxyalkanoate (PHA) granules related to PHB. Notably, DsRed2EC-LactC2 expressed in Magnetospirillum gryphiswaldense was detected at the position of membrane-enclosed magnetosome chains and at the cytoplasmic membrane but not at PHB granules. In conclusion, the carbonosomes of representatives of α-proteobacteria, β-proteobacteria and γ-proteobacteria have no phospholipids in vivo and we postulate that the PHB/PHA granule surface layers in natural producers generally are free of phospholipids and consist of proteins only.

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Smart polyhydroxyalkanoate nanobeads by protein based functionalization

Cited by 53 publications

References 137 publications

Polyhydroxyalkanoate‐associated phasins as phylogenetically heterogeneous, multipurpose proteins

Polyhydroxyalkanoate‐associated phasins as phylogenetically heterogeneous, multipurpose proteins

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

Polyhydroxyalkanoate (PHA) Granules Have no Phospholipids

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