scholarly journals Engineering of Fatty Acid Synthases (FASs) to Boost the Production of Medium-Chain Fatty Acids (MCFAs) in Mucor circinelloides

2019 ◽  
Vol 20 (3) ◽  
pp. 786 ◽  
Author(s):  
Syed Hussain ◽  
Ahsan Hameed ◽  
Md. Khan ◽  
Yao Zhang ◽  
Huaiyuan Zhang ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
De Novo ◽  
Acyl Carrier Protein ◽  
Fold Increase ◽  
Mucor Circinelloides ◽  
Palm Tree ◽  
Medium Chain ◽  
Health Related ◽  
Fatty Acid Synthases

Increasing energy demands and health-related concerns worldwide have motivated researchers to adopt diverse strategies to improve medium-chain fatty acid (MCFA) biosynthesis for use in the functional food and aviation industries. The abundance of naturally produced MCFAs from botanical sources (i.e., coconut fruit/seeds and palm tree) has been observed to be insufficient compared with the various microorganisms used to cope with industrial demands. Mucor circinelloides is one of many promising microorganisms; it exhibits diverse biotechnological importance ranging from the production of functional lipids to applications in the manufacture of bio-fuel. Thus, research was conducted to acquire the desired elevated amounts of MCFAs (i.e., C8–C12) from metabolically engineered strains of M. circinelloides M65. To achieve this goal, four different acyl-acyl carrier protein (ACP) thioesterase (TE)-encoding genes exhibiting a substrate preference for medium-chain acyl-ACP molecules were expressed in M. circinelloides M65, resulting in the generation of C8–C12 fatty acids. Among all the engineered strains, M65-TE-03 and M65-TE-04 demonstrated the highest production of non-native C8–C10 and C12 fatty acids, respectively, in comparison to the control. These recombinant strains biosynthesized MCFAs de novo within the range from 28 to 46% (i.e., 1.14 to 2.77 g/L) of total cell lipids. Moreover, the reduction in chain length eventually resulted in a 1.5–1.75-fold increase in total lipid productivity in the engineered strains. The MCFAs were also found to be integrated into all lipid classes. This work illustrates how the integration of heterologous enzymes in M. circinelloides can offer a novel opportunity to edit the fatty acid synthases (FAS) complex, resulting in increased production of microbial MFCAs.

scholarly journals Molecular cloning and characterisation of an acyl carrier protein thioesterase gene (CocoFatB1) expressed in the endosperm of coconut (Cocos nucifera) and its heterologous expression in Nicotiana tabacum to engineer the accumulation of different fatty acids

2014 ◽  
Vol 41 (1) ◽  
pp. 80 ◽  
Author(s):  
Yijun Yuan ◽  
Yinhua Chen ◽  
Shan Yan ◽  
Yuanxue Liang ◽  
Yusheng Zheng ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Amino Acid ◽  
Nicotiana Tabacum ◽  
Acyl Carrier Protein ◽  
Cocos Nucifera ◽  
Chain Elongation ◽  
Carrier Protein ◽  
Medium Chain ◽  
Tobacco Seeds

Coconut (Cocos nucifera L.) contains large amounts of medium chain fatty acids, which mostly recognise acyl-acyl carrier protein (ACP) thioesterases that hydrolyse acyl-ACP into free fatty acids to terminate acyl chain elongation during fatty acid biosynthesis. A full-length cDNA of an acyl-ACP thioesterase, designated CocoFatB1, was isolated from cDNA libraries prepared from coconut endosperm during fruit development. The gene contained an open reading frame of 1254 bp, encoding a 417-amino acid protein. The amino acid sequence of the CocoFatB1 protein showed 100% and 95% sequence similarity to CnFatB1 and oil palm (Elaeis guineensis Jacq.) acyl-ACP thioesterases, respectively. Real-time fluorescent quantitative PCR analysis indicated that the CocoFatB1 transcript was most abundant in the endosperm from 8-month-old coconuts; the leaves and endosperm from 15-month-old coconuts had ~80% and ~10% of this level. The CocoFatB1 coding region was overexpressed in tobacco (Nicotiana tabacum L.) under the control of the seed-specific napin promoter following Agrobacterium tumefaciens-mediated transformation. CocoFatB1 transcript expression varied 20-fold between different transgenic plants, with 21 plants exhibiting detectable levels of CocoFatB1 expression. Analysis of the fatty acid composition of transgenic tobacco seeds showed that the levels of myristic acid (14 : 0), palmitic acid (16 : 0) and stearic acid (18 : 0) were increased by 25%, 34% and 17%, respectively, compared with untransformed plants. These results indicated that CocoFatB1 acts specifically on 14 : 0-ACP, 16 : 0-ACP and 18 : 0-ACP, and can increase medium chain saturated fatty acids. The gene may valuable for engineering fatty acid metabolism in crop improvement programmes.

scholarly journals Triacylglycerol synthesis in goat mammary gland. The effect of ATP, Mg2+ and glycerol 3-phosphate on the esterification of fatty acids synthesized de novo

1984 ◽  
Vol 220 (2) ◽  
pp. 513-519 ◽  
Author(s):  
H O Hansen ◽  
I Grunnet ◽  
J Knudsen
Keyword(s):  
Fatty Acids ◽  
Mammary Gland ◽  
Fatty Acid ◽  
De Novo ◽  
Fatty Acid Synthetase ◽  
Acid Synthesis ◽  
Medium Chain ◽  
Medium Chain Fatty Acids ◽  
Triacylglycerol Synthesis ◽  
Chain Fatty Acids

Goat mammary-gland microsomal fraction by itself induces synthesis of medium-chain-length fatty acids by goat mammary fatty acid synthetase and incorporates short- and medium-chain fatty acids into triacylglycerol. Addition of ATP in the absence or presence of Mg2+ totally inhibits triacylglycerol synthesis from short- and medium-chain fatty acids, and severely inhibits synthesis de novo of medium-chain fatty acids. The inhibition by ATP of fatty acid synthesis and triacylglycerol synthesis de novo can be relieved by glycerol 3-phosphate. The effect of ATP could not be mimicked by the non-hydrolysable ATP analogue, adenosine 5′-[beta, gamma-methylene]triphosphate and could not be shown to be caused by inhibition of the diacylglycerol acyltransferase by a phosphorylation reaction. Possible explanations for the mechanism of the inhibition by ATP are discussed, and a hypothetical model for its action is outlined.

scholarly journals Leptin controls the fate of fatty acids in isolated rat white adipocytes

2002 ◽  
Vol 175 (3) ◽  
pp. 735-744 ◽  
Author(s):  
◽  
RB Ceddia ◽  
R Curi
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Free Fatty Acids ◽  
De Novo ◽  
Fold Increase ◽  
Acid Synthesis ◽  
Exogenous Glucose ◽  
White Adipocytes ◽  
Direct Energy ◽  
Isolated Rat

Leptin directly increases the rate of exogenous glucose and fatty acids oxidation in isolated adipocytes. However, the effects of leptin on fatty acid metabolism in white adipose tIssue have not been examined in detail. Here, we report that in adipocytes incubated for 6 h in the presence of leptin (10 ng/ml), the insulin-stimulated de novo fatty acid synthesis was inhibited by 36% (P<0.05), while the exogenous oxidation of acetic and oleic acids was increased by 50% and 76% respectively. Interestingly, leptin did not alter the oxidation of intracellular fatty acids. Leptin-incubated cells presented a 16-fold increase in the incorporation of oleic acid into triglyceride (TG) and a 123% increase in the intracellular TG hydrolysis (as measured by free fatty acids release). Fatty acid-TG cycling was not affected by leptin. By employing fatty acids radiolabeled with (3)H and (14)C, we could determine the concomitant influx of fatty acids (incorporation of fatty acids into TG) and efflux of fatty acids (intracellular fatty acids oxidation and free fatty acids release) in the incubated cells. Leptin increased by 30% the net efflux of fatty acids from adipocytes. We conclude that leptin directly inhibits de novo synthesis of fatty acids and increases the release and oxidation of fatty acids in isolated rat adipocytes. These direct energy-dissipating effects of leptin may play an important role in reducing accumulation of fatty acids into TG of rat adipose cells.

scholarly journals The fractionation of the fatty acid synthetase activities of avocado mesocarp plastids

1975 ◽  
Vol 146 (2) ◽  
pp. 439-445 ◽  
Author(s):  
P J Weaire ◽  
R G O Kekwick
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
De Novo ◽  
Acyl Carrier Protein ◽  
Fatty Acid Synthetase ◽  
Carrier Protein ◽  
Acetyl Coa ◽  
Principal Product ◽  
Absolute Requirement ◽  
Malonyl Coa

1. The range of fatty acids formed by preparations of ultrasonically ruptured avocado mesocarp plastids was dependent on the substrate. Whereas [1-14C]palmitate and [14C]oleate were the major products obtained from [-14C]acetate and [1-14C]acetyl-CoA, the principal product from [2-14C]malonyl-CoA was [14-C]stearate. 2. Ultracentrifugation of the ruptured plastids at 105000g gave a supernatant that formed mainly stearate from [2-14C]malonyl-CoA and to a lesser extent from [1-14C]acetate. The incorporation of [1-14C]acetate into stearate by this fraction was inhibited by avidin. 3. The 105000g precipitate of the disrupted plastids incorporated [1-14C]acetate into a mixture of fatty acids that contained largely [14C]plamitate and [14C]oleate. The formation of [14C]palmitate and [14C]oleate by disrupted plastids was unaffected by avidin. 4. The soluble fatty acid synthetase was precipitated from the 105000g supernatant in the 35-65%-saturated-(NH4)2SO4 fraction and showed an absolute requirement for acyl-carrier protein. 5. Both fractions synthesized fatty acids de novo.

scholarly journals Insights into triterpene synthesis and unsaturated fatty-acid accumulation provided by chromosomal-level genome analysis of Akebia trifoliata subsp. australis

2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Hui Huang ◽  
Juan Liang ◽  
Qi Tan ◽  
Linfeng Ou ◽  
Xiaolin Li ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Unsaturated Fatty Acids ◽  
Developmental Stages ◽  
De Novo ◽  
Acyl Carrier Protein ◽  
Repetitive Sequences ◽  
Gene Families ◽  
Terpene Synthase ◽  
Comparative Genomic

AbstractAkebia trifoliata subsp. australis is a well-known medicinal and potential woody oil plant in China. The limited genetic information available for A. trifoliata subsp. australis has hindered its exploitation. Here, a high-quality chromosome-level genome sequence of A. trifoliata subsp. australis is reported. The de novo genome assembly of 682.14 Mb was generated with a scaffold N50 of 43.11 Mb. The genome includes 25,598 protein-coding genes, and 71.18% (485.55 Mb) of the assembled sequences were identified as repetitive sequences. An ongoing massive burst of long terminal repeat (LTR) insertions, which occurred ~1.0 million years ago, has contributed a large proportion of LTRs in the genome of A. trifoliata subsp. australis. Phylogenetic analysis shows that A. trifoliata subsp. australis is closely related to Aquilegia coerulea and forms a clade with Papaver somniferum and Nelumbo nucifera, which supports the well-established hypothesis of a close relationship between basal eudicot species. The expansion of UDP-glucoronosyl and UDP-glucosyl transferase gene families and β-amyrin synthase-like genes and the exclusive contraction of terpene synthase gene families may be responsible for the abundant oleanane-type triterpenoids in A. trifoliata subsp. australis. Furthermore, the acyl-ACP desaturase gene family, including 12 stearoyl-acyl-carrier protein desaturase (SAD) genes, has expanded exclusively. A combined transcriptome and fatty-acid analysis of seeds at five developmental stages revealed that homologs of SADs, acyl-lipid desaturase omega fatty acid desaturases (FADs), and oleosins were highly expressed, consistent with the rapid increase in the content of fatty acids, especially unsaturated fatty acids. The genomic sequences of A. trifoliata subsp. australis will be a valuable resource for comparative genomic analyses and molecular breeding.

The role of acyl carrier protein isoforms from Cuphea lanceolata seeds in the de-novo biosynthesis of medium-chain fatty acids

Planta ◽  
1998 ◽  
Vol 205 (2) ◽  
pp. 263-268 ◽  
Author(s):  
Burkhardt Siegfried Schütt ◽  
Monika Brummel ◽  
Ricardo Schuch ◽  
Friedrich Spener
Keyword(s):  
Fatty Acids ◽  
De Novo ◽  
Acyl Carrier Protein ◽  
Protein Isoforms ◽  
Carrier Protein ◽  
Medium Chain ◽  
Medium Chain Fatty Acids ◽  
De Novo Biosynthesis ◽  
Cuphea Lanceolata

scholarly journals Optimization of Diverse Carbon Sources and Cultivation Conditions for Enhanced Growth and Lipid and Medium-Chain Fatty Acid (MCFA) Production by Mucor circinelloides

Fermentation ◽  
2019 ◽  
Vol 5 (2) ◽  
pp. 35 ◽  
Author(s):  
Hussain ◽  
Nazir ◽  
Hameed ◽  
Yang ◽  
Mustafa ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Carbon Sources ◽  
Mucor Circinelloides ◽  
Chain Fatty Acid ◽  
Medium Chain Fatty Acid ◽  
Microbial Lipids ◽  
Cultivation Conditions ◽  
Medium Chain ◽  
Culture Optimization

Keywords: Mucor circinelloides; microbial lipids; medium-chain fatty acids; culture optimization

scholarly journals Chimeric Fatty Acyl-Acyl Carrier Protein Thioesterases Provide Mechanistic Insight into Enzyme Specificity and Expression

2018 ◽  
Vol 84 (10) ◽  
Author(s):  
Marika Ziesack ◽  
Nathan Rollins ◽  
Aashna Shah ◽  
Brendon Dusel ◽  
Gordon Webster ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Acyl Carrier Protein ◽  
Fatty Acyl ◽  
Carrier Protein ◽  
Fatty Acid Production ◽  
Medium Chain ◽  
Commodity Chemicals ◽  
Long Chain ◽  
Chain Fatty Acids

ABSTRACT Medium-chain fatty acids are commodity chemicals. Increasing and modifying the activity of thioesterases (TEs) on medium-chain fatty acyl-acyl carrier protein (acyl-ACP) esters may enable a high-yield microbial production of these molecules. The plant Cuphea palustris harbors two distinct TEs: C. palustris FatB1 ( Cp FatB1) (C 8 specificity, lower activity) and Cp FatB2 (C 14 specificity, higher activity) with 78% sequence identity. We combined structural features from these two enzymes to create several chimeric TEs, some of which showed nonnatural fatty acid production as measured by an enzymatic assay and gas chromatography-mass spectrometry (GC-MS). Notably, chimera 4 exhibited an increased C 8 fatty acid production in correlation with improved microbial expression. This chimera led us to identify Cp FatB2-specific amino acids between positions 219 and 272 that lead to higher protein levels. Chimera 7 produced a broad range of fatty acids and appeared to combine a fatty acid binding pocket with long-chain specificity and an ACP interaction site that may activate fatty acid extrusion. Using homology modeling and in silico docking with ACP, we identified a “positive patch” within amino acids 162 to 218, which may direct the ACP interaction and regulate access to short-chain fatty acids. On the basis of this modeling, we transplanted putative ACP interaction sequences from Cp FatB1 into Cp FatB2 and created a chimeric thioesterase that produced medium-chain as well as long-chain fatty acids. Thus, the engineering of chimeric enzymes and characterizing their microbial activity and chain-length specificity suggested mechanistic insights into TE functions and also generated thioesterases with potentially useful properties. These observations may inform a rational engineering of TEs to allow alkyl chain length control. IMPORTANCE Medium-chain fatty acids are important commodity chemicals. These molecules are used as plastic precursors and in shampoos and other detergents and could be used as biofuel precursors if production economics were favorable. Hydrocarbon-based liquid fuels must be optimized to have a desired boiling point, low freezing point, low viscosity, and other physical characteristics. Similarly, the solubility and harshness of detergents and the flexibility of plastic polymers can be modulated. The length and distribution of the carbon chains in the hydrophobic tails determine these properties. The biological synthesis of cell membranes and fatty acids produces chains of primarily 16 to 18 carbons, which give rise to current biofuels. The ultimate goal of the work presented here is to engineer metabolic pathways to produce designer molecules with the correct number of carbons in a chain, so that such molecules could be used directly as specialty commodity chemicals or as fuels after minimal processing.

Sign in / Sign up

Export Citation Format

Share Document