scholarly journals Engineering the Monomer Composition of Polyhydroxyalkanoates Synthesized in Saccharomyces cerevisiae

2006 ◽  
Vol 72 (1) ◽  
pp. 536-543 ◽  
Author(s):  
Bo Zhang ◽  
Ross Carlson ◽  
Friedrich Srienc
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Fatty Acid ◽  
Chain Length ◽  
Fatty Acid Metabolism ◽  
Acid Metabolism ◽  
Pha Synthase ◽  
Fatty Acid Degradation ◽  
Medium Chain ◽  
Acid Degradation ◽  
Wide Range

ABSTRACT Polyhydroxyalkanoates (PHAs) have received considerable interest as renewable-resource-based, biodegradable, and biocompatible plastics with a wide range of potential applications. We have engineered the synthesis of PHA polymers composed of monomers ranging from 4 to 14 carbon atoms in either the cytosol or the peroxisome of Saccharomyces cerevisiae by harnessing intermediates of fatty acid metabolism. Cytosolic PHA production was supported by establishing in the cytosol critical β-oxidation chemistries which are found natively in peroxisomes. This platform was utilized to supply medium-chain (C6 to C14) PHA precursors from both fatty acid degradation and synthesis to a cytosolically expressed medium-chain-length (mcl) polymerase from Pseudomonas oleovorans. Synthesis of short-chain-length PHAs (scl-PHAs) was established in the peroxisome of a wild-type yeast strain by targeting the Ralstonia eutropha scl polymerase to the peroxisome. This strain, harboring a peroxisomally targeted scl-PHA synthase, accumulated PHA up to approximately 7% of its cell dry weight. These results indicate (i) that S. cerevisiae expressing a cytosolic mcl-PHA polymerase or a peroxisomal scl-PHA synthase can use the 3-hydroxyacyl coenzyme A intermediates from fatty acid metabolism to synthesize PHAs and (ii) that fatty acid degradation is also possible in the cytosol as β-oxidation might not be confined only to the peroxisomes. Polymers of even-numbered, odd-numbered, or a combination of even- and odd-numbered monomers can be controlled by feeding the appropriate substrates. This ability should permit the rational design and synthesis of polymers with desired material properties.

scholarly journals Identification and Characterization of a New Enoyl Coenzyme A Hydratase Involved in Biosynthesis of Medium-Chain-Length Polyhydroxyalkanoates in Recombinant Escherichia coli

2003 ◽  
Vol 185 (18) ◽  
pp. 5391-5397 ◽  
Author(s):  
Si Jae Park ◽  
Sang Yup Lee
Keyword(s):  
Escherichia Coli ◽  
Fatty Acid ◽  
Chain Length ◽  
Biosynthetic Pathway ◽  
Pha Synthase ◽  
Enzymatic Analysis ◽  
E Coli ◽  
Medium Chain ◽  
Medium Chain Length ◽  
Enoyl Coa Hydratase

ABSTRACT The biosynthetic pathway of medium-chain-length (MCL) polyhydroxyalkanoates (PHAs) from fatty acids has been established in fadB mutant Escherichia coli strain by expressing the MCL-PHA synthase gene. However, the enzymes that are responsible for the generation of (R)-3-hydroxyacyl coenzyme A (R3HA-CoAs), the substrates for PHA synthase, have not been thoroughly elucidated. Escherichia coli MaoC, which is homologous to Pseudomonas aeruginosa (R)-specific enoyl-CoA hydratase (PhaJ1), was identified and found to be important for PHA biosynthesis in a fadB mutant E. coli strain. When the MCL-PHA synthase gene was introduced, the fadB maoC double-mutant E. coli WB108, which is a derivative of E. coli W3110, accumulated 43% less amount of MCL-PHA from fatty acid compared with the fadB mutant E. coli WB101. The PHA biosynthetic capacity could be restored by plasmid-based expression of the maoCEc gene in E. coli WB108. Also, E. coli W3110 possessing fully functional β-oxidation pathway could produce MCL-PHA from fatty acid by the coexpression of the maoCEc gene and the MCL-PHA synthase gene. For the enzymatic analysis, MaoC fused with His6-Tag at its C-terminal was expressed in E. coli and purified. Enzymatic analysis of tagged MaoC showed that MaoC has enoyl-CoA hydratase activity toward crotonyl-CoA. These results suggest that MaoC is a new enoyl-CoA hydratase involved in supplying (R)-3-hydroxyacyl-CoA from the β-oxidation pathway to PHA biosynthetic pathway in the fadB mutant E. coli strain.

scholarly journals Reply: “Letter to the Editor Re: Billeaud et al. Nutrients 2018, 10, 690”

Nutrients ◽  
2019 ◽  
Vol 11 (2) ◽  
pp. 406
Author(s):  
Claude Billeaud ◽  
Carole Boué-Vaysse ◽  
Leslie Couëdelo ◽  
Philippe Steenhout ◽  
Jonathan Jaeger ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Docosahexaenoic Acid ◽  
Human Milk ◽  
Preterm Infants ◽  
Fatty Acid Metabolism ◽  
Acid Metabolism ◽  
Medium Chain ◽  
Letter To The Editor ◽  
Human Milk Fortifier

We thank Bernard and colleagues for their careful reading and interest in our article Effects on Fatty Acid Metabolism of a New Powdered Human Milk Fortifier Containing Medium-Chain Triacylglycerols and Docosahexaenoic Acid in Preterm Infants [...]

scholarly journals Bacillus subtilis Gene Cluster Involved in Calcium Carbonate Biomineralization

2006 ◽  
Vol 189 (1) ◽  
pp. 228-235 ◽  
Author(s):  
Chiara Barabesi ◽  
Alessandro Galizzi ◽  
Giorgio Mastromei ◽  
Mila Rossi ◽  
Elena Tamburini ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Fatty Acid ◽  
Calcium Carbonate ◽  
Fatty Acid Metabolism ◽  
Insertional Mutagenesis ◽  
Molecular Mechanisms ◽  
Acid Metabolism ◽  
Precipitation Process ◽  
Calcium Carbonate Precipitation ◽  
Wide Range

ABSTRACT Calcium carbonate precipitation, a widespread phenomenon among bacteria, has been investigated due to its wide range of scientific and technological implications. Nevertheless, little is known of the molecular mechanisms by which bacteria foster calcium carbonate mineralization. In our laboratory, we are studying calcite formation by Bacillus subtilis, in order to identify genes involved in the biomineralization process. A previous screening of UV mutants and of more than one thousand mutants obtained from the European B. subtilis Functional Analysis project allowed us to isolate strains altered in the precipitation phenotype. Starting from these results, we focused our attention on a cluster of five genes (lcfA, ysiA, ysiB, etfB, and etfA) called the lcfA operon. By insertional mutagenesis, mutant strains carrying each of the five genes were produced. All of them, with the exception of the strain carrying the mutated lcfA operon, were unable to form calcite crystals. By placing transcription under IPTG (isopropyl-β-d-thiogalactopyranoside) control, the last gene, etfA, was identified as essential for the precipitation process. To verify cotranscription in the lcfA operon, reverse transcription-PCR experiments were performed and overlapping retrocotranscripts were found comprising three adjacent genes. The genes have putative functions linked to fatty acid metabolism. A link between calcium precipitation and fatty acid metabolism is suggested.

scholarly journals Identification and characterization of an acyl-CoA dehydrogenase from Pseudomonas putida KT2440 that shows preference towards medium to long chain length fatty acids

Microbiology ◽  
2014 ◽  
Vol 160 (8) ◽  
pp. 1760-1771 ◽  
Author(s):  
Maciej W. Guzik ◽  
Tanja Narancic ◽  
Tatjana Ilic-Tomic ◽  
Sandra Vojnovic ◽  
Shane T. Kenny ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Pseudomonas Putida ◽  
Chain Length ◽  
Recombinant Expression ◽  
Nutrient Utilization ◽  
Ecological Niches ◽  
Fatty Acid Degradation ◽  
Acid Degradation

Diverse and elaborate pathways for nutrient utilization, as well as mechanisms to combat unfavourable nutrient conditions make Pseudomonas putida KT2440 a versatile micro-organism able to occupy a range of ecological niches. The fatty acid degradation pathway of P. putida is complex and correlated with biopolymer medium chain length polyhydroxyalkanoate (mcl-PHA) biosynthesis. Little is known about the second step of fatty acid degradation (β-oxidation) in this strain. In silico analysis of its genome sequence revealed 21 putative acyl-CoA dehydrogenases (ACADs), four of which were functionally characterized through mutagenesis studies. Four mutants with insertionally inactivated ACADs (PP_1893, PP_2039, PP_2048 and PP_2437) grew and accumulated mcl-PHA on a range of fatty acids as the sole source of carbon and energy. Their ability to grow and accumulate biopolymer was differentially negatively affected on various fatty acids, in comparison to the wild-type strain. Inactive PP_2437 exhibited a pattern of reduced growth and PHA accumulation when fatty acids with lengths of 10 to 14 carbon chains were used as substrates. Recombinant expression and biochemical characterization of the purified protein allowed functional annotation in P. putida KT2440 as an ACAD showing clear preference for dodecanoyl-CoA ester as a substrate and optimum activity at 30 °C and pH 6.5–7.

Fatty acid metabolism in Saccharomyces cerevisiae

2003 ◽  
Vol 60 (9) ◽  
pp. 1838-1851 ◽  
Author(s):  
C. W. T. van Roermund ◽  
H. R. Waterham ◽  
L. Ijlst ◽  
R. J. A. Wanders
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Fatty Acid ◽  
Fatty Acid Metabolism ◽  
Acid Metabolism

scholarly journals Effects on Fatty Acid Metabolism of a New Powdered Human Milk Fortifier Containing Medium-Chain Triacylglycerols and Docosahexaenoic Acid in Preterm Infants

Nutrients ◽  
2018 ◽  
Vol 10 (6) ◽  
pp. 690 ◽  
Author(s):  
Claude Billeaud ◽  
Carole Boué-Vaysse ◽  
Leslie Couëdelo ◽  
Philippe Steenhout ◽  
Jonathan Jaeger ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Docosahexaenoic Acid ◽  
Human Milk ◽  
Preterm Infants ◽  
Fatty Acid Metabolism ◽  
Acid Metabolism ◽  
Medium Chain ◽  
Human Milk Fortifier

scholarly journals A Label-Free Proteomic Strategy to Investigate the Intramuscular fat Proteomic Differences Between Biceps Femoris and Longissimus Dorsi in Inner Mongolian Cashmere Goats

2020 ◽  
Author(s):  
Yuchun Xie ◽  
Zhihong Liu ◽  
Rile Nai ◽  
Juntao Guo ◽  
Xin Su ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Fatty Acid Metabolism ◽  
Acid Metabolism ◽  
Biceps Femoris ◽  
Intramuscular Fat ◽  
Longissimus Dorsi ◽  
Fatty Acid Elongation ◽  
Label Free ◽  
Fatty Acid Degradation ◽  
Cashmere Goats

Abstract Background: As a major raw-cashmere-producing province in China. Nearly 700,000 Aerbasi cashmere goats are fed per year, and the corresponding meat production is nearly 10,000 tons. However, there are no reports on the meat of this goat. To better understand the molecular variations underlying intramuscular fat (IMF) anabolism and catabolism in Inner Mongolian cashmere goats, the proteomic differences between the biceps femoris (BF) and longissimus dorsi (LD) were investigated by a label-free strategy. Then, the identified proteins were verified as being involved in IMF anabolism and catabolism by Western blot analysis.Results: The IMF content was significantly higher in the BF than in the LD, suggesting that IMF accumulated more in the BF or was metabolized more in the LD. We performed proteomic analysis of IMF anabolism and catabolism at the proteomic level, and 1209 proteins were identified in the BF (high-IMF) and LD (low-IMF) groups. Among them, 110 were differentially expressed proteins (DEPs), 81 of which were upregulated in the high-IMF group, while 29 were upregulated in the low-IMF group. Gene ontology (GO) classification showed that the 110 DEPs were functionally classified into 100 annotation clusters. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that the 110 DEPs covered 34 KEGG pathways. Three pathways were related to IMF metabolism and deposition—fatty acid metabolism, fatty acid degradation and fatty acid elongation—and included 7 proteins.Conclusion: GO and KEGG analyses showed that differentially expressed HADHA, HADHB, ACSL1, ACADS, ACAT1 and ACAA2 in the mitochondria act via fatty acid metabolism, fatty acid degradation and fatty acid elongation to influence the metabolism and synthesis of long-, short- and medium-chain fatty acids and modulate IMF anabolism and catabolism. Protein-protein interaction (PPI) network analysis showed that IMF accumulation in different muscle tissues of Inner Mongolian cashmere goats was affected not only by 5 key enzymes and proteins involved in fatty acid synthesis and metabolism but also by five DEPs (SUCLG1, SUCLG2, CS, DLST, and ACO2) in the TCA cycle. Our results provide new insights into IMF deposition in goats and improve our understanding of the molecular mechanisms underlying IMF anabolism and catabolism.

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