Non-biodegradable biopolymers from renewable resources: perspectives and impacts.

In recent years the biotechnological production of bulk biopolymers has focused on the synthesis of biodegradable polymers to replace their non-biodegradable counterparts derived from fossil resources. Examples include polyhydroxyalkanoates and polylactic acid, which act as substitutes for polyolefins. By contrast, the biotechnological production of non-biodegradable polymers from renewable resources has so far been scarcely considered, probably because this idea contradicts the paradigm that all natural compounds are biodegradable.

Establishment of Tn5096-based transposon mutagenesis in Gordonia polyisoprenivorans.

The transposons Tn5, Tn10, Tn611, and Tn5096 were characterized regarding transposition in Gordonia polyisoprenivorans strain VH2. No insertional mutants were obtained employing Tn5 or Tn10. The thermosensitive plasmid pCG79 harboring Tn611 integrated into the chromosome of G.

Occurrence and expression of tricarboxylate synthases in Ralstonia eutropha.

2-Methylcitrate synthase (2-MCS1) and citrate synthase (CS) of Ralstonia eutropha strain H16 were separated by affinity chromatography and analyzed for their substrate specificities. 2-MCS1 used not only the primary substrate propionyl-CoA but also acetyl-CoA and, at a low rate, even butyryl-CoA and valeryl-CoA for condensation with oxaloacetate. The KM values for propionyl-CoA and acetyl-CoA were 0.

Degradation of cyanophycin by Sedimentibacter hongkongensis strain KI and Citrobacter amalonaticus strain G Isolated from an anaerobic bacterial consortium.

Using a combination of various enrichment techniques, the strictly anaerobic, gram-positive, endospore-forming bacterium Sedimentibacter hongkongensis strain KI as revealed by 16S rRNA analysis and the gram-negative enterobacterium Citrobacter amalonaticus strain G as revealed by physiological tests were isolated from an anaerobic cyanophycin (CGP)-degrading bacterial consortium. S. hongkongensis strain KI is the first anaerobic bacterium with the ability to hydrolyze CGP to beta-Asp-Arg and beta-Asp-Lys dipeptides, as revealed by electrospray ionization-mass spectrometry and reversed-phase high-performance liquid chromatography analysis.

Biosynthesis of poly (3-mercaptopropionate) and poly (3-mercaptopropionate-co-3-hydroxybutyrate) with recombinant Escherichia coli.

Polythioesters newly emerged as a type of novel polymer and they have showed great potential for application in industries. In this study, genes of butyrate kinase (buk) and phosphotransbutyrylase (ptb) from Clostridium acetobutylicum, and poly (3-hydroxybutyrate) (PHB) synthase gene from Thiocapsa pfennigii were used for construction of a metabolic pathway to synthesize the polythioesters. When 3-mercaptopropionate and 3-hydroxybutyrate were fed, poly (3-mercaptopropoinate) [poly (3MP)] and poly(3-mercaptopropionate-co-3-hydroxybutyrate) [poly(3MP-co-3HB)] were synthesized by recombinant Escherichia coli JM109 (pBPP1) harboring the constructed metabolic pathway.

Investigations on the solubility behavior of cyanophycin. Solubility of cyanophycin in solutions of simple inorganic salts.

On the basis of a previous report on the occurrence of water-soluble cyanophycin (CGP, cyanophycin granule polypeptide) in a recombinant strain of Escherichia coli expressing the cyanophycin synthetase (CphA) of Desulfitobacterium hafniense published by others, the conditions of its production were investigated in this study. Although the incubation temperature, aeration level, and NaCl concentration during cultivation had effects on the in vivo production of water-soluble CGP, it could be isolated as a major variant irrespective of the cultivation conditions. The occurrence of the soluble variant was also not dependent on the E.

Gordonia nitida Yoon et al. 2000 is a later synonym of Gordonia alkanivorans Kummer et al. 1999.

The name of the species Gordonia nitida is validly published but its type strain DSM 44499(T) shares high similarity based on 16S rRNA gene sequences with Gordonia alkanivorans DSM 44369(T) and Gordonia westfalica DSM 44215(T). These three species obviously build up a distinct cluster within the genus Gordonia. In the present paper, data from the literature concerning the three Gordonia species were reviewed and the genetic similarity of G.

Poly(3-mercaptopropionate): a nonbiodegradable biopolymer?

Polythioesters (PTEs) represent a novel class of biopolymers, which basically can be synthesized with polyhydroxyalkanoate (PHA) biosynthesis systems. Albeit technical applications of PTEs have not been elucidated yet, biodegradability might be an important property of this new thermoplastic material. In this study, extensive approaches were employed to isolate microorganisms capable of degrading poly(3-mercaptopropionate), poly(3MP), as a model compound of PTEs.

Poly(3-hydroxybutyrate) granule-associated proteins: impacts on poly(3-hydroxybutyrate) synthesis and degradation.

Polyhydroxyalkanoates (PHAs) represent a group of biopolymers that are synthesized by many bacteria as storage compounds and deposited as insoluble cytoplasmic inclusions. Because they have many putative technical and medical applications, PHAs may play an important role in human life in the future. Therefore, for academic interest the bacterial PHA metabolism has been studied in much detail.

Influence of homologous phasins (PhaP) on PHA accumulation and regulation of their expression by the transcriptional repressor PhaR in Ralstonia eutropha H16.

Phasins play an important role in the formation of poly(3-hydroxybutyrate) [poly(3HB)] granules and affect their size. Recently, three homologues of the phasin protein PhaP1 were identified in Ralstonia eutropha strain H16. The functions of PhaP2, PhaP3 and PhaP4 were examined by analysis of R.

Physiological conditions conducive to high cyanophycin content in biomass of Acinetobacter calcoaceticus strain ADP1.

The effects of the inorganic medium components, the initial pH, the incubation temperature, the oxygen supply, the carbon-to-nitrogen ratio, and chloramphenicol on the synthesis of cyanophycin (CGP) by Acinetobacter calcoaceticus strain ADP1 were studied in a mineral salts medium containing sodium glutamate and ammonium sulfate as carbon and nitrogen sources, respectively. Variation of all these factors resulted in maximum CGP contents of only about 3.5% (wt/wt) of the cell dry matter (CDM), and phosphate depletion triggered CGP accumulation most substantially.

Application of the BPEC pathway for large-scale biotechnological production of poly(3-mercaptopropionate) by recombinant Escherichia coli, including a novel in situ isolation method.

Metabolically engineered Escherichia coli JM109 harboring plasmid pBPP1 and expressing the nonnatural BPEC pathway for synthesis of thermoplastic polyhydroxyalkanoates (PHA) and novel polythioesters (PTE) to provide suitable substrates of PHA synthase was investigated with respect to biotechnological production of poly(3-mercaptopropionate) [poly(3MP)]. Fed-batch fermentation processes were established at the 30- and 500-liter scales in stirred tank bioreactors to produce kilogram amounts of poly(3MP). Cultivation was done in a modified M9 mineral salts medium containing glucose or glycerol as the carbon and energy source and with 3-mercaptopropionic acid (3MP) as the precursor substrate for poly(3MP) biosynthesis provided from the late exponential growth phase.

Thio wax ester biosynthesis utilizing the unspecific bifunctional wax ester synthase/acyl coenzyme A:diacylglycerol acyltransferase of Acinetobacter sp. strain ADP1.

The bifunctional wax ester synthase/acyl coenzyme A (acyl-CoA):diacylglycerol acyltransferase (WS/DGAT) from Acinetobacter sp. strain ADP1 (formerly Acinetobacter calcoaceticus ADP1) mediating the biosyntheses of wax esters and triacylglycerols was used for the in vivo and in vitro biosynthesis of thio wax esters and dithio wax esters. For in vitro biosynthesis, 5'His(6)WS/DGAT comprising an N-terminal His(6) tag was purified from the soluble protein fraction of Escherichia coli Rosetta(DE3)pLysS (pET23a::5'His(6)atf).

The wax ester synthase/acyl coenzyme A:diacylglycerol acyltransferase from Acinetobacter sp. strain ADP1: characterization of a novel type of acyltransferase.

The wax ester synthase/acyl coenzyme A (acyl-CoA):diacylglycerol acyltransferase (WS/DGAT) catalyzes the final steps in triacylglycerol (TAG) and wax ester (WE) biosynthesis in the gram-negative bacterium Acinetobacter sp. strain ADP1. It constitutes a novel class of acyltransferases which is fundamentally different from acyltransferases involved in TAG and WE synthesis in eukaryotes.

Mechanism of lipid-body formation in prokaryotes: how bacteria fatten up.

Neutral lipid accumulation is frequently observed in some Gram-negative prokaryotes like Acinetobacter sp. and most actinomycetes, including the pathogenic Mycobacterium tuberculosis and antibiotic producing streptomycetes. We examined the formation of wax ester- and triacylglycerol (TAG)-bodies in Acinetobacter calcoaceticus and Rhodococcus opacus using microscopic, immunological and biophysical methods.

Identification and characterization of genes from Streptomyces sp. strain K30 responsible for clear zone formation on natural rubber latex and poly(cis-1,4-isoprene) rubber degradation.

Streptomyces sp. strain K30 was isolated from soil next to a city high way in Münster (Germany) according to its ability to degrade natural and synthetic poly(cis-1,4-isoprene) rubber and to form clear zones on natural rubber latex agar plates. The clear zone forming phenotype was used to clone the responsible gene by phenotypic complementation of a clear zone negative mutant.

Synthesis of novel lipids in Saccharomyces cerevisiae by heterologous expression of an unspecific bacterial acyltransferase.

The bifunctional wax ester synthase/acyl-coenzyme A:diacylglycerol acyltransferase (WS/DGAT) is the key enzyme in storage lipid accumulation in the gram-negative bacterium Acinetobacter calcoaceticus ADP1, mediating wax ester, and to a lesser extent, triacylglycerol (TAG) biosynthesis. Saccharomyces cerevisiae accumulates TAGs and steryl esters as storage lipids. Four genes encoding a DGAT (Dga1p), a phospholipid:diacylglycerol acyltransferase (Lro1p) and two acyl-coenzyme A:sterol acyltransferases (ASATs) (Are1p and Are2p) are involved in the final esterification steps in TAG and steryl ester biosynthesis in this yeast.

Caenibacterium thermophilum is a later synonym of Schlegelella thermodepolymerans.

Recently, two strains of Schlegelella thermodepolymerans Elbanna et al. 2003 and an independently isolated bacterium, Caenibacterium thermophilum Manaia et al. 2003, were described in parallel as gen.

Physiological and morphological responses of the soil bacterium Rhodococcus opacus strain PD630 to water stress.

Rhodococcus opacus PD630 was investigated for physiological and morphological changes under water stress challenge. Gluconate- and hexadecane-grown cells were extremely resistant to these conditions, and survival accounted for up to 300 and 400 days; respectively, when they were subjected to slow air-drying. Results of this study suggest that strain PD630 has specific mechanisms to withstand water stress.

Microbial polythioesters.

This feature article describes the current knowledge on biosynthesis of polythioesters (PTEs), which are exclusively accumulated by microorganisms capable to synthesize the well-known polyhydroxyalkanoates (PHAs). Two bacterial PTE biosynthesis systems are discussed, both are depending on the cultivation conditions and appropriate feeding regimes. The first system comprises the production of PTE copolymers by Ralstonia eutropha, and the second system has been established in recombinant Escherichia coli to produce PTE homopolymers.

Studies on the biodegradability of polythioester copolymers and homopolymers by polyhydroxyalkanoate (PHA)-degrading bacteria and PHA depolymerases.

The biodegradability of microbial polythioesters (PTEs), a novel class of biopolymers which were discovered recently and can be produced by polyhydroxyalkanoate (PHA)-accumulating bacteria, was studied. Using poly(3-hydroxybutyrate- co-3-mercaptopropionate) [poly(3HB- co-3MP)] as sole carbon source for screening, 22 new bacterial strains were isolated and characterized. Interestingly, none of the PHA-degrading bacteria was able to utilize the homopolymer poly(3MP) as a carbon source for growth or to form clear zones on poly(3MP)-containing agar plates.

Polyhydroxyalkanoate (PHA) accumulation in sulfate-reducing bacteria and identification of a class III PHA synthase (PhaEC) in Desulfococcus multivorans.

Seven strains of sulfate-reducing bacteria (SRB) were tested for the accumulation of polyhydroxyalkanoates (PHAs). During growth with benzoate Desulfonema magnum accumulated large amounts of poly(3-hydroxybutyrate) [poly(3HB)]. Desulfosarcina variabilis (during growth with benzoate), Desulfobotulus sapovorans (during growth with caproate), and Desulfobacterium autotrophicum (during growth with caproate) accumulated poly(3HB) that accounted for 20 to 43% of cell dry matter.

Partial purification and characterization of a non-cyanobacterial cyanophycin synthetase from Acinetobacter calcoaceticus strain ADP1 with regard to substrate specificity, substrate affinity and binding to cyanophycin.

This study reports, for the first time, purification and biochemical characterization of a cyanophycin synthetase from a non-cyanobacterial strain. Cyanophycin synthetase of Acinetobacter calcoaceticus strain ADP1 was purified 69-fold from recombinant Escherichia coli by two chromatographic steps and one novel affinity step utilizing the Mg(2+)-dependent binding of the enzyme to cyanophycin. Unlike cyanobacterial cyanophycin synthetases characterized so far, the purified enzyme from A.