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1.
Keenan TM Nakas JP Tanenbaum SW 《Journal of industrial microbiology & biotechnology》2006,33(7):616-626
The potential for the use of woody biomass in poly-β-hydroxyalkanoate (PHA) biosynthesis is reviewed. Based on previously cited work indicating incorporation of xylose or levulinic acid (LA) into PHAs by several bacterial strains, we have initiated a study for exploring bioconversion of forest resources to technically relevant copolymers. Initially, PHA was synthesized in shake-flask cultures of Burkholderia cepacia grown on 2.2% (w/v) xylose, periodically amended with varying concentrations of levulinic acid [0.07–0.67% (w/v)]. Yields of poly(β-hydroxybutyrate-co-β-hydroxyvalerate) [P(3HB-co-3HV)] from 1.3 to 4.2 g/l were obtained and could be modulated to contain from 1.0 to 61 mol% 3-hydroxyvalerate (3HV), as determined by 1H and 13C NMR analyses. No evidence for either the 3HB or 4HV monomers was found. Characterization of these P(3HB-co-3HV) samples, which ranged in molecular mass (viscometric, M
v) from 511–919 kDa, by differential scanning calorimetry and thermogravimetric analyses (TGA) provided data which were in agreement for previously reported P(3HB-co-3HV) copolymers. For these samples, it was noted that melting temperature (T
m) and glass transition temperature (T
g) decreased as a function of 3HVcontent, with T
m demonstrating a pseudoeutectic profile as a function of mol% 3HV content. In order to extend these findings to the use of hemicellulosic process streams as an inexpensive carbon source, a detoxification procedure involving sequential overliming and activated charcoal treatments was developed. Two such detoxified process hydrolysates (NREL CF: aspen and CESF: maple) were each fermented with appropriate LA supplementation. For the NREL CF hydrolysate-based cultures amended with 0.25–0.5% LA, P(3HB-co-3HV) yields, PHA contents (PHA as percent of dry biomass), and mol% 3HV compositions of 2.0 g/l, 40% (w/w), and 16–52 mol% were obtained, respectively. Similarly, the CESF hydrolysate-based shake-flask cultures yielded 1.6 g/l PHA, 39% (w/w) PHA contents, and 4–67 mol% 3HV compositions. These data are comparable to copolymer yields and cellular contents reported for hexose plus levulinic acid-based shake-flask cultures, as reported using Alcaligenes eutrophus and Pseudomonas putida. However, our findings presage a conceivable alternative, forestry-based biorefinery approach for the production of value-added biodegradable PHA polymers. Specifically, this review describes the current and potential utilization of lignocellulosic process streams as platform precursors to PHA polymers including hemicellulosic hydrolysates, residual cellulose-derived levulinic acid, tall oil fatty acids (Kraft pulping residual), and lignin-derived aromatics. 相似文献
2.
Solaiman DK 《Journal of industrial microbiology & biotechnology》2003,30(5):322-326
Pseudomonas resinovorans harbors two isogenic poly(hydroxyalkanoates) (PHAs) synthase genes (phaC1
Pre
, phaC2
Pre
) responsible for the production of intracellular medium-chain-length (mcl-)PHAs. Sequence analysis showed that the putative
gene-products of these genes contain a conserved α/β-hydrolase fold in the carboxy-terminal half of the proteins. Hybrid genes pha7 and pha8 were constructed by exchanging the α/β-hydrolase-fold coding portions of phaC1
Pre
and phaC2
Pre
at the 3′ terminal. When grown with decanoate as carbon source, the pha7- or pha8-transformed Escherichia coli LS1298 produced PHAs containing 73–75% β-hydroxydecanoate (β-HD) and 25–27% β-hydroxyoctanoate (β-HO). Deletion mutants,
Δpha7 and Δpha8, were isolated during the PCR-based construction of pha7 and pha8, respectively. Cells harboring these mutants produced PHAs containing 55–60 mol% β-HD and 40–45 mol% β-HO. These results
demonstrate the feasibility of generating active hybrid mcl-PHA synthase genes and their mutants with the potential of producing
polymers having a varied repeat-unit composition. 相似文献
3.
Polyhydroxyalkanoates (PHAs) are biological polyesters, of which, Short-Chain-Length-Medium-Chain-Length (SCL-MCL) PHA copolymers
are important because of their wide range of applications. The present study focused on molecular characterization of Pseudomonas sp. LDC-5 that is identified as SCL-MCL producer. Phase contrast, fluorescent and electron microscopic observation confirmed
the presence of PHA granules in Pseudomonas sp. LDC-5. PCR analysis indicated the presence of expected amplicon for SCL phaC gene (∼500 bp), MCL phaC1 with phaZ (∼1.3), and phaC2 with phaZ (∼1.5 kb). Sequence analysis of the PHA synthase gene of Pseudomonas sp. LDC-5 revealed significant differences in phaC1 and phaC2 which were further confirmed by recombinant studies. Recombinant Escherichia coli harboring the partial phaC1 gene was able to accumulate PHA, whereas E. coli with phaC2 did not accumulate PHA as verified by fold analysis, immunoblotting, Gas Chromatography (GC), Differential scanning calorimetry
(DSC), and FTIR studies. The predicted theoretical three-dimensional structure revealed that PhaC1 is consistent with α/β hydrolase fold. Monomer
composition showed the presence of monomer ranging from C4 to C12: 1 when glucose and sodium octanoate fed as the carbon source.
DSC revealed melting temperature peak at 153.12°C and glass transition (Tg) peaks at −0.37°C. Thermogravimetric analysis revealed that the polymer was stable up to 276°C. Fourier Transform Infrared
Spectroscopy (FT-IR) spectral analysis showed the PHA specific wave number at 1,739.67 and 1,161.07 cm−1. The potential of Pseudomonas sp. LDC-5 and its properties are discussed. 相似文献
4.
Recombinant strains of Wautersia eutropha expressing an artificial polyhydroxyalkanoate (PHA) biosynthesis operon under the control of different native promoters linked to polyhydroxybutyrate (PHB) (Pphb), acetoin (PacoE, PacoD, and PacoX) or pyruvate (PpdhE) metabolism were constructed and tested. The promoters were representative either of the enterobacterial σ70 (Pphb, PacoE, and PpdhE)- or σ54 (PacoD and PacoX)-dependent promoters. To obtain polymers consisting of C4–C12 monomer units, an artificial operon consisting of the PHA synthase gene from Pseudomonas sp. 61-3 (phaC1
Ps) tandemly linked to the W. eutropha genes encoding β-ketothiolase (phbA
We) and nicotinamide adenine dinucleotide phosphate dependent acetoacetyl-coenzyme A (CoA) reductase (phbB
We) was constructed. All recombinant strains produced PHA, indicating that the PHA biosynthesis genes were expressed under the control of the different promoters. Cell growth and PHA synthesis on MS medium complemented with gluconate or octanoate, and different concentrations of acetoin (0, 0.15, and 0.3%) clearly differed among the recombinant strains. While the PacoD and PacoX promoters mediated only low PHA yields (<1%) in the presence of the inducer acetoin, the remaining promoters—independent of the addition of acetoin—resulted in the production of PHA polymers with high 3HB fractions (90–100 mol%) and with high 3HO contents (70–86 mol%) from gluconate and octanoate, respectively. Interestingly, on octanoate-MS medium with 0.15% acetoin, the PacoE promoter mediated the synthesis of PHA with a relatively high 3HB fraction (48 mol%). While PHAs with high 3HB contents were obtained, the overall PHA product yields were low (<10%); thus, their potential application for further commercial exploitation appears limited. 相似文献
5.
Rodrigues MF Valentin HE Berger PA Tran M Asrar J Gruys KJ Steinbüchel A 《Applied microbiology and biotechnology》2000,53(4):453-460
Burkholderia sp. accumulates polyhydroxyalkanoates (PHAs) containing 3-hydroxybutyrate and 3-hydroxy-4-pentenoic acid when grown on mineral
media under limited phosphate or nitrogen, and using sucrose or gluconate as a carbon and energy source. Solvent fractionation
and NMR spectroscopic characterization of these polyesters revealed the simultaneous accumulation of two homopolyesters rather
than a co-polyester with random sequence distribution of the monomers [Valentin HE, Berger PA, Gruys KJ, Rodrigues MFA, Steinbüchel
A, Tran M, Asrar J (1999) Macromolecules 32: 7389–7395]. To understand the genetic requirements for such unusual polyester
accumulation, we probed total genomic DNA from Burkholderia sp. by Southern hybridization experiments using phaC-specific probes. These experiments indicated the presence of more than one PHA synthase gene within the genome of Burkholderia sp. However, when total genomic DNA from Burkholderia sp. was used to complement a PHA-negative mutant of Ralstonia eutropha for PHA accumulation, only one PHA synthase gene was obtained resembling the R. eutropha type of PHA synthases, based on amino acid sequence similarity. In addition to the PHA synthase gene, based on high sequence
homology, genes encoding a β-ketothiolase and acetoacetyl-CoA reductase were identified in a gene cluster with the PHA synthase
gene. The arrangement of the three genes is quite similar to the R. eutropha poly-β-hydroxybutyrate biosynthesis operon.
Received: 3 September 1999 / Received revision: 29 October 1999 / Accepted: 5 November 1999 相似文献
6.
Synthesis of poly(3-hydroxyalkanoates) by mutant and recombinant Pseudomonas strains 总被引:1,自引:0,他引:1
Gjalt W. Huisman Eric Wonink Gertjan de Koning Hans Preusting Bernard Witholt 《Applied microbiology and biotechnology》1992,38(1):1-5
We have studied the accumulation kinetics and physical characteristics of the poly(3-hydroxyalkanoates) (PHAs) formed by several Pseudomonas strains, mutants and recombinants. Although PHA synthesis generally begins only after an essential nutrient such as N, P, S or Mg becomes limiting, we have identified at least one strain (P. putida KT2442) that begins producing PHA during the exponential growth phase. This PHA is chemically and physically identical to that produced by P. oleovorans GPol, the strain in which we first identified PHA. Analysis of the PHA formed by a mutant strain defective in PHA degradation (P. oleovorans GPo500) revealed that the molecular mass (Mw), the monomer composition and thermal characteristics were similar to that of the PHA of the wild-type parent strain P. oleovorans GPo1. The pha locus of P. oleovorans encodes enzymes that are involved in PHA biosynthesis and degradation. It has been subcloned to study the two PHA polymerases separately in a PHA– mutant (GPp104) derived from P. putida KT2442. The recombinant strains accumulated lower PHA levels than the wild-type strains, and the Mw of these polymers were lower than those produced by the wild-type P. oleovorans and parent strain. The monomer composition of the two PHAs formed by the two PHA polymerases differed, indicating that the PHA polymerases have different substrate specificities for the incorporation of 3-hydroxyoctanoate and 3-hydroxyhexanoate monomers into PHA. Despite these differences, the PHAs formed were essentially indistinguishable from wild-type PHAs with respect to their thermal characteristics.Correspondence to: B. Witholt 相似文献
7.
Habibah A Wahab Nurul Bahiyah Ahmad Khairudin Mohd Razip Samian Nazalan Najimudin 《BMC structural biology》2006,6(1):23-14
Background
Polyhydroxyalkanoates (PHA), are biodegradable polyesters derived from many microorganisms such as the pseudomonads. These polyesters are in great demand especially in the packaging industries, the medical line as well as the paint industries. The enzyme responsible in catalyzing the formation of PHA is PHA synthase. Due to the limited structural information, its functional properties including catalysis are lacking. Therefore, this study seeks to investigate the structural properties as well as its catalytic mechanism by predicting the three-dimensional (3D) model of the Type II Pseudomonas sp. USM 4–55 PHA synthase 1 (PhaC1P.sp USM 4–55). 相似文献8.
Volova TG Trusova MY Kalacheva GS Kozhevnicov IV 《Applied microbiology and biotechnology》2006,73(2):429-433
Physiological–biochemical, genetic, and cultural properties of the glucose-utilizing mutant strain Ralstonia eutropha B8562 have been compared with those of its parent strain R. eutropha B5786. It has been shown that growth characteristics of the strain cultured on glucose as the sole carbon and energy source are comparable with those of the parent strain. Strain B8562 is characterized by high polyhydroxyalkanoate (PHA) yields on different carbon sources (CO2, fructose, and glucose). PHA accumulation in the strain batch cultured on glucose under nitrogen deficiency reaches 90 %. The major monomer in the PHA is β-hydroxybutyric acid (more than 99 mol %); the identified minor components are β-hydroxyvaleric acid (0.25–0.72 mol %) and β-hydroxyhexanoic acid (0.08–1.5 mol %). The strain is a promising PHA producer on available sugar-containing media with glucose. 相似文献
9.
Kathryn L. Houmiel Steven Slater Debra Broyles Laura Casagrande Susan Colburn Kathleen Gonzalez Timothy A. Mitsky Steven E. Reiser Devang Shah Nancy B. Taylor Mintien Tran Henry E. Valentin Kenneth J. Gruys 《Planta》1999,209(4):547-550
Polyhydroxyalkanoates (PHAs) comprise a class of biodegradable polymers which offer an environmentally sustainable alternative
to petroleum-based plastics. Production of PHAs in plants is attractive since current fermentation technology is prohibitively
expensive. The PHA homopolymer poly(β-hydroxybutyrate) (PHB) has previously been produced in leaves of Arabidopsis thaliana (Nawrath et al., 1994, Proc Natl Acad Sci USA 91: 12760–12764). However, Brassica napus oilseed may provide a better system for PHB production because acetyl-CoA, the substrate required in the first step of PHB
biosynthesis, is prevalent during fatty acid biosynthesis. Three enzymatic activities are needed to synthesize PHB: a β-ketothiolase,
an acetoacetyl-CoA reductase and a PHB synthase. Genes from the bacterium Ralstonia eutropha encoding these enzymes were independently engineered behind the seed-specific Lesquerella fendleri oleate 12-hydroxylase promoter in a modular fashion. The gene cassettes were sequentially transferred into a single, multi-gene
vector which was used to transform B. napus. Poly(β-hydroxybutyrate) accumulated in leukoplasts to levels as high as 7.7% fresh seed weight of mature seeds. Electron-microscopy
analyses indicated that leukoplasts from these plants were distorted, yet intact, and appeared to expand in response to polymer
accumulation.
Received: 26 May 1999 / Accepted: 16 June 1999 相似文献
10.
Pengtao Huang Takaya Okoshi Shoji Mizuno Ayaka Hiroe 《Bioscience, biotechnology, and biochemistry》2018,82(9):1615-1623
Medium-chain-length (mcl)-polyhydroxyalkanoates (PHAs), elastomeric polyesters synthesized by Genus Pseudomonas bacteria, generally have many different monomer components. In this study, PHAs biosynthesized by four type strains of Pseudomonas (P. putida, P. citronellolis, P. oleovorans, and P. pseudoalcaligenes) and a typical PHA producer (P. putida KT2440) were characterized in terms of the monomer structure and composition by gas chromatography-mass spectrometry (GC-MS) analysis. With a thiomethyl pretreatment of PHA methanolysis derivatives, two unsaturated monomers, 3-hydroxy-5-dodecenoate (3H5DD) and 3-hydroxy-5-tetradecenoate (3H5TD), were identified in mcl-PHAs produced by P. putida and P. citronellolis. The quantitative analysis of PHA monomers was performed by employing GC-MS with methanolysis derivatives, and the results coincided with those obtained by performing nuclear magnetic resonance spectroscopy. Only poly(3-hydroxybutyrate) was detected from the P. oleovorans and P. pseudoalcaligenes type strains. These analytical results would be useful as a reference standard for phenotyping of new PHA-producing bacteria. 相似文献
11.
Carmen Scholz 《Applied microbiology and biotechnology》2010,88(4):829-837
An overview is provided on the possibilities of producing positively and negatively charged poly(β-hydroxyalkanoates), PHAs.
A large variety of bacterial polyesters with functionalized terminal side chains can be produced in microbial fermentation
processes by a direct polymerization of respective carbon sources, that is, carbon sources that carry functional groups in
their ω-position. However, charged PHAs are not accessible by a direct approach and must be synthesized via polymer-analogous
reactions of functionalized bacterial polyesters. PHA polyanions are produced by converting the terminal functional groups
into carboxylate groups, while PHA polycations are produced by introducing terminal amino groups. PHAs with terminal vinyl
groups emerged as most suitable PHA precursors, as they can be produced in relatively high yields and the double bonds are
sufficiently reactive. The oxidation of vinyl groups yields PHA polyanions. The conversion of terminal vinyl groups into epoxides
with a subsequent ring-opening reaction with an amine yields PHA polycations. Other functionalized PHA that potentially lend
themselves to polymer-analogous reactions are reviewed. 相似文献
12.
Michael Knoll Thomas M Hamm Florian Wagner Virginia Martinez Jürgen Pleiss 《BMC bioinformatics》2009,10(1):89
Background
Polyhydroxyalkanoates (PHAs) can be degraded by many microorganisms using intra- or extracellular PHA depolymerases. PHA depolymerases are very diverse in sequence and substrate specificity, but share a common α/β-hydrolase fold and a catalytic triad, which is also found in other α/β-hydrolases. 相似文献13.
Molecular characterization based on 16s rDNA gene sequence analysis of bacterial colonies isolated from endosulfan contaminated
soil showed the presence of Ochrobacterum sp, Burkholderia sp, Pseudomonas alcaligenes, Pseudomonas sp and Arthrobacter sp which degraded 57–90% of α-endosulfan and 74–94% of β-endosulfan after 7days. Whole cells of Pseudomonas sp and Pseudomonas alcaligenes showed 94 and 89% uptake of α-isomer and 86 and 89% of β-endosulfan respectively in 120 min. In Pseudomonas sp, endosulfan sulfate was the major metabolite detected during the degradation of α-isomer, with minor amount of endosulfan
diol while in Pseudomonas alcaligenes endosulfan diol was the only product during α-endosulfan degradation. Whole cells of Pseudomonas sp also utilized 83% of endosulfan sulfate in 120 min. In situ applications of the defined consortium consisting of Pseudomonas alcaligenes and Pseudomonas sp (1:1) in plots contaminated with endosulfan showed that 80% of α-endosulfan and 65% of β-endosulfan was degraded after
12 weeks of incubation. Endosulfan sulfate formed during endosulfan degradation was subsequently degraded to unknown metabolites.
ERIC-PCR analysis indicated 80% survival of introduced population of Pseudomonas alcaligenes and Pseudomonas sp in treated plots. 相似文献
14.
Polyhydroxyalkanoates (PHAs) are biodegradable polymers produced by microbes to overcome environmental stress. Commercial
production of PHAs is limited by the high cost of production compared to conventional plastics. Another hindrance is the brittle
nature and low strength of polyhydroxybutyrate (PHB), the most widely studied PHA. The needs are to produce PHAs, which have
better elastomeric properties suitable for biomedical applications, preferably from inexpensive renewable sources to reduce
cost. Certain unique properties of Bacillus subtilis such as lack of the toxic lipo-polysaccharides, expression of self-lysing genes on completion of PHA biosynthetic process
– for easy and timely recovery, usage of biowastes as feed enable it to compete as potential candidate for commercial production
of PHA. 相似文献
15.
Matsusaki H Abe H Taguchi K Fukui T Doi Y 《Applied microbiology and biotechnology》2000,53(4):401-409
Pseudomonas sp. 61-3 accumulated a blend of poly(3-hydroxybutyrate) [P(3HB)] homopolymer and a random copolymer consisting of 3-hydroxyalkanoate
(3HA) units of 4–12 carbon atoms. The genes encoding β-ketothiolase (PhbARe) and NADPH-dependent acetoacetyl-CoA reductase (PhbBRe) from Ralstoniaeutropha were expressed under the control of promoters for Pseudomonas sp. 61-3 pha locus or R. eutropha phb operon together with phaC1
Ps gene (PHA synthase 1 gene) from Pseudomonas sp. 61-3 in PHA-negative mutants P. putida GPp104 and R. eutropha PHB−4 to produce copolyesters [P(3HB-co-3HA)] consisting of 3HB and medium-chain-length 3HA units of 6–12 carbon atoms. The introduction of the three genes into
GPp104 strain conferred the ability to synthesize P(3HB-co-3HA) with relatively high 3HB compositions (up to 49 mol%) from gluconate and alkanoates, although 3HB units were not incorporated
at all or at a very low fraction (3 mol%) into copolyesters by the strain carrying phaC1
Ps gene only. In addition, recombinant strains of R. eutropha PHB−4 produced P(3HB-co-3HA) with higher 3HB fractions from alkanoates and plant oils than those from recombinant GPp104 strains. One of the recombinant
strains, R. eutropha PHB−4/pJKSc46-pha, in which all the genes introduced were expressed under the control of the native promoter for Pseudomonas sp. 61-3 pha locus, accumulated P(3HB-co-3HA) copolyester with a very high 3HB fraction (85 mol%) from palm oil. The nuclear magnetic resonance analyses showed that
the copolyesters obtained here were random copolymers of 3HB and 3HA units.
Received: 12 July 1999 / Received revision: 1 October 1999 / Accepted: 2 October 1999 相似文献
16.
We investigated the expression of (R)-specific enoyl coenzyme A hydratase (PhaJ) in Pseudomonas putida KT2440 accumulating polyhydroxyalkanoate (PHA) from sodium octanoate in order to identify biosynthesis pathways of PHAs from
fatty acids in pseudomonads. From a database search through the P. putida KT2440 genome, an additional phaJ gene homologous to phaJ4
Pa from Pseudomonas aeruginosa, termed phaJ4
Pp, was identified. The gene products of phaJ1
Pp, which was identified previously, and phaJ4
Pp were confirmed to be functional in recombinant Escherichia coli on PHA synthesis from sodium dodecanoate. Cytosolic proteins from P. putida grown on sodium octanoate were subjected to anion exchange chromatography and one of the eluted fractions with hydratase
activity included PhaJ4Pp, as revealed by western blot analysis. These results strongly suggest that PhaJ4Pp forms a channeling route from β-oxidation to PHA biosynthesis in P. putida. Moreover, the substrate specificity of PhaJ1Pp was suggested to be different from that of PhaJ1Pa from P. aeruginosa although these two proteins share 67% amino acid sequence identity. 相似文献
17.
Phosphobacteria are able to enhance phosphorus availability in soil and improve crop yields. To develop such biofertilizers,
14 predominant phosphobacteria were isolated from eutrophic aquatic ecosystems. Molecular identification and phylogenetic
analysis revealed three groups among the nine isolates of inorganic phosphate-solubilizing bacteria (IPSB): IPSBl and IPSB2
belonged to the actinobacteria and flavobacteria, respectively, and the other seven belonged to the γ-proteobacteria. Among
five isolates of organic phosphorus-mineralizing bacteria (OPMB), two groups were present: OPMB1 and OPMB3 belonged to the
β-proteobacteria, while the other three belonged to the γ-proteobacteria. The IPSB isolates released 62.8–66.7 mg P 1−1 from tricalcium phosphate under shaking conditions, and 26.8 to 43.7 mg P 1−1 under static conditions; the OPMB strains released 23.5–30.2 mg P 1−1 from lecithin under shaking conditions, and 16.7–27.6 mg P 1−1 under static conditions. To the best of our knowledge, this is the first report indicating that IPSBl (designated Aureobacterium resistents) as a tricalcium phosphate-solubilizing bacterium and OPMB1 and OPMB3 (designated Acidovorax temperans and Achromobacter xylosoxidans, respectively) are lecithin-mineralizing bacteria. This investigation demonstrated that a eutrophic aquatic ecosystem is
a selective source of phosphobacteria and the screened phosphobacteria are a potential alternative to the development of biofertilizers. 相似文献
18.
The time course of the accumulation of triacylglycerols (TAGs) in Rhodococcus opacus PD630 or of TAGs plus polyhydroxyalkanoates (PHA) in Rhodococcus ruber NCIMB 40126 with gluconate or glucose as carbon source, respectively, was studied. In addition, we examined the mobilization
of these storage compounds in the absence of a carbon source. R. opacus accumulated TAGs only after the exhaustion of ammonium in the medium, and, with a fixed concentration of the carbon source,
the amounts of TAGs in the cells increased with decreasing concentrations of ammonium in the medium. When these cells were
incubated in the absence of an additional carbon source, about 90% of these TAGs were mobilized and used as endogenous carbon
source, particularly if ammonium was available. R. ruber accumulated a copolyester consisting of 3-hydroxybutyrate and 3-hydroxyvalerate already during the early exponential growth
phase, whereas TAGs were synthesized and accumulated mainly during the late exponential and stationary growth phases. In the
stationary growth phase, synthesis of TAGs continued, whereas PHA was partially mobilized. In the absence of an additional
carbon source but in the presence of ammonium, mobilization of TAGs started first and was then paralleled by the mobilization
of PHA, resulting in an approximately 90% and 80% decrease of these storage compounds, respectively. During the accumulation
phase, interesting shifts in the composition of the two storage compounds occurred, indicating that the substrates of the
PHA synthase and the TAG synthesizing enzymes were provided to varying extents, depending on whether the cells were in the
early or late exponential or in the stationary growth phase.
Received: 12 January 2000 / Received revision: 22 February 2000 / Accepted: 25 February 2000 相似文献
19.
Four (R)-specific enoyl CoA hydratases (PhaJ) interconnect the β-oxidation pathway with PHA biosynthesis in Pseudomonas aeruginosa. The use of antisense technique and over-expression to delineate the role of two of these enzymes, PhaJ1 and PhaJ4 forms
the basis of this study. It has been observed that P. aeruginosa recombinant with phaJ1 antisense construct, fed with different fatty acids, produces PHA with less hydroxy octanoate (7–11% reduction) and a proportionate
increase in other monomer fractions, compared to that of the control. Recombinants bearing phaJ4 antisense construct are found to contain less hydroxy decanoate (10–11% reduction) and more or less equal amount of hydroxy
octanoate, compared to that of the control. P. aeruginosa has produced PHA with more hydroxy octanoate and decanoate (6–17% increase), respectively, when PhaJ1 and PhaJ4 have been
over-expressed individually or along with PhaC1. PhaJ1 and PhaJ4 are found to be involved mainly in the production of hydroxy
octanoyl CoA and hydroxy decanoyl CoA, respectively, in P. aeruginosa. The strongest accumulation of hydroxy octanoate and hydroxy decanoate has been observed along with hydroxy butyrate, in
PHA, produced by E. coli, when PhaC1 has been co-expressed with PhaJ1 and PhaJ4, respectively. We have demonstrated, for the first time, the polymerization
of hydroxy butyryl CoA monomers in recombinant E. coli by PhaC1 of P. aeruginosa. 相似文献
20.
A polyhydroxyalkanoate (PHA) synthase gene phaC2
Ps from Pseudomonas stutzeri strain 1317 was introduced into a PHA synthase gene phbC
Re negative mutant, Ralstonia eutropha PHB−4. It conferred on the host strain the ability to synthesize PHA, the monomer compositions of which varied widely when grown
on different carbon sources. During cultivation on gluconate, the presence of phaC2
Ps in R. eutropha PHB−4 led to the accumulation of polyhydroxybutyrate (PHB) homopolymer in an amount of 40.9 wt% in dry cells. With fatty acids,
the recombinant successfully produced PHA copolyesters containing both short-chain-length and medium-chain-length 3-hydroxyalkanoate
(3HA) of 4–12 carbon atoms in length. When cultivated on a mixture of gluconate and fatty acid, the monomer composition of
accumulated PHA was greatly affected and the monomer content was easily regulated by the addition of fatty acids in the cultivation
medium. After the (R)-3-hydroxydecanol-ACP:CoA transacylase gene phaG
Pp from Pseudomonas putida was introduced into phaC2
Ps-containing R. eutropha PHB−4, poly(3HB-co-3HA) copolyester with a very high 3-hydroxybutyrate (3HB) fraction (97.3 mol%) was produced from gluconate and the monomer
compositions of PHA synthesized from fatty acids were also altered. This study clearly demonstrated that PhaC2Ps cloned from P. stutzeri 1317 has extraordinarily low substrate specificity in vivo, though it has only 54% identity in comparison to a previously described low-substrate-specificity PHA synthase PhaC1Ps from Pseudomonas sp. 61–3. This study also indicated that the monomer composition and content of the synthesized PHA can be effectively modulated
by controlling the addition of carbon sources or by modifying metabolic pathways in the hosts. 相似文献