Cloning and analysis of the polyhydroxyalkanoic acid synthase gene from an Acinetobacter sp.: Evidence that the gene is both plasmid and chromosomally located

Schembri, Mark A.

doi:10.1016/0378-1097(94)90610-6

Cited by 12 publications

(24 citation statements)

References 16 publications

(26 reference statements)

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“…PHB accumulation has been shown to be induced by phosphate starvation in all Acinetobacter PHB-positive strains examined in our laboratory (31). The regulation of PHB biosynthesis has been studied extensively in A. eutrophus (33).…”

Section: Discussionmentioning

confidence: 90%

“…The Acinetobacter PHA synthase gene (phaC Ac ) was cloned by heterologous complementation in the PHB-negative mutant A. eutrophus PHB-4 as previously described (31). Nucleotide sequencing both upstream and downstream of phaC Ac on pJKD1226 revealed the presence of two other PHA biosynthetic genes, phaB Ac (encoding an acetoacetyl-CoA reductase) and phaA Ac (encoding a ␤-ketothiolase) (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…3). The Acinetobacter phaC Ac gene product has previously been characterized and shows significant similarity to other PHA synthase proteins (31).…”

Section: Resultsmentioning

confidence: 99%

“…A previous report has shown that the Acinetobacter PHA synthase gene (phaC Ac ) may be present in more than one copy (31). Therefore, it is possible that the primer extension products observed upstream of ORF1 in fact result from a second copy of this gene within the Acinetobacter genome.…”

Section: Discussionmentioning

confidence: 99%

“…Recently, the PHA synthase gene was cloned from an Acinetobacter strain isolated from a modified activated sludge treatment plant (31). Acinetobacter species have been suggested to be the major organisms responsible for enhanced biological phosphate removal from wastewater treated in alternating anaerobic/aerobic activated sludge systems (5).…”

mentioning

confidence: 99%

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Phosphate concentration regulates transcription of the Acinetobacter polyhydroxyalkanoic acid biosynthetic genes

1995

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The polyhydroxyalkanoic acid (PHA) biosynthetic gene locus was cloned and characterized from an Acinetobacter sp. isolated from activated sludge. Nucleotide sequence analysis identified three clustered genes, phaA Ac (encoding a ␤-ketothiolase), phaB Ac (encoding an acetoacetyl coenzyme A reductase), and phaC Ac (encoding a PHA synthase). In addition, an open reading frame (ORF1) with potential to encode a 13-kDa protein was identified within this locus. The sequence of the putative translational product of ORF1 does not show significant similarity to any sequences in the database. A plasmid containing the Acinetobacter pha locus conferred the ability to accumulate poly-␤-hydroxybutyrate on its Escherichia coli host. These genes appear to lie in an operon transcribed by two promoters upstream of phaB Ac , an apparent constitutive promoter, and a second promoter induced by phosphate starvation and under pho regulon control. These as well as a number of additional potential transcription start points were identified by a combination of primer extension and promoter-chloramphenicol acetyltransferase gene fusion studies carried out in Acinetobacter or E. coli transformants.Polyhydroxyalkanoic acids (PHAs) are a family of bacterial polyesters synthesized by a wide range of organisms under conditions of nutrient limitation in the presence of an excess carbon and energy source (1). These polymers have attracted significant attention for their potential commercial exploitation as biodegradable plastics (22). The most abundant and best-studied PHA is poly-␤-hydroxybutyrate (PHB), a linear, unbranched homopolymer built up of (R)-3-hydroxybutyric acid units.The physiology and molecular genetics of PHB biosynthesis have been extensively studied in Alcaligenes eutrophus (33). More recently, genes involved in PHA metabolism have also been cloned from numerous other organisms (see the review in reference 32). In most organisms, PHB is synthesized via a three-step pathway which involves the condensation of two molecules of acetyl coenzyme A (acetyl-CoA) to acetoacetylCoA via a ␤-ketothiolase (PhaA), reduction of acetoacetylCoA to D-(Ϫ)-␤-hydroxybutyryl-CoA via an NADPH-dependent acetoacetyl-CoA reductase (PhaB), and polymerization to PHB via a PHA synthase (PhaC) (33).Recently, the PHA synthase gene was cloned from an Acinetobacter strain isolated from a modified activated sludge treatment plant (31). Acinetobacter species have been suggested to be the major organisms responsible for enhanced biological phosphate removal from wastewater treated in alternating anaerobic/aerobic activated sludge systems (5). Biochemical models of this overall process involve the breakdown of polyphosphate and synthesis of PHB during the anaerobic stage and breakdown of PHB and production of polyphosphate in the subsequent aerobic stage (8). Strains of Acinetobacter which can accumulate up to 10% phosphorus (10) and 15% PHB (28) on a dry weight basis have been isolated.In A. eutrophus, all three PHA biosynthetic enzymes are synthesized constitutiv...

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Section: Discussionmentioning

confidence: 90%

Section: Resultsmentioning

confidence: 99%

“…3). The Acinetobacter phaC Ac gene product has previously been characterized and shows significant similarity to other PHA synthase proteins (31).…”

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Phosphate concentration regulates transcription of the Acinetobacter polyhydroxyalkanoic acid biosynthetic genes

1995

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PHB and Other Polhydroxyalkanoic Acids

Steinbüchel¹

1996

Biotechnology

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Effect of the applied organic load rate on biodegradable polymer production by mixed microbial cultures in a sequencing batch reactor

Dionisi

Majone

Vallini

et al. 2005

Biotech & Bioengineering

168

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This article studies the operation of a new process for the production of biopolymers (polyhydroxyalkanoates, PHAs) at different applied organic load rates (OLRs). The process is based on the aerobic enrichment of activated sludge to obtain mixed cultures able to store PHAs at high rates and yields. A mixture of acetic, lactic, and propionic acids at different concentrations (in the range 8.5-31.25 gCOD/L) was fed every 2 h in a sequencing batch reactor (SBR). The resulting applied OLR was in the range 8.5-31.25 gCOD/L/day. Even though, as expected, the increase in the OLR caused an increase in biomass concentration (up to about 8.7 g COD/L), it also caused a relevant decrease of maximal polymer production rate. This decrease in polymer production rate was related to the different extent of "feast and famine" conditions, as function of the applied OLR and of the start-up conditions. As a consequence the best performance of the process was obtained at an intermediate OLR (20 gCOD/L/day) where both biomass productivity and PHA storage were high enough. However, at this high OLR the process was unstable and sudden decrease of performance was also observed. The sludge characterized by the highest PHA storage response was investigated by 16S rDNA clone library. The clone library contained sequences mostly from PHA producers (e.g., Alcaligenes and Comamonas genera); however many genera and among them, one of the dominant (Thauera), were never described before in relation to PHA storage response.

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Cloning and analysis of the polyhydroxyalkanoic acid synthase gene from an Acinetobacter sp.: Evidence that the gene is both plasmid and chromosomally located

Cited by 12 publications

References 16 publications

Phosphate concentration regulates transcription of the Acinetobacter polyhydroxyalkanoic acid biosynthetic genes

Phosphate concentration regulates transcription of the Acinetobacter polyhydroxyalkanoic acid biosynthetic genes

PHB and Other Polhydroxyalkanoic Acids

Effect of the applied organic load rate on biodegradable polymer production by mixed microbial cultures in a sequencing batch reactor

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