The Experts below are selected from a list of 309 Experts worldwide ranked by ideXlab platform
Randall J. Weselake - One of the best experts on this subject based on the ideXlab platform.
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Diacylglycerol Acyltransferase 1 is activated by phosphatidate and inhibited by SnRK1‐catalyzed phosphorylation
Plant Journal, 2018Co-Authors: Kristian Mark P. Caldo, Randall J. Weselake, Wei Shen, Yang Xu, Linda Hanley-bowdoin, Guanqun Chen, M. Joanne LemieuxAbstract:Diacylglycerol Acyltransferase 1 (DGAT1) catalyzes the final and committed step in the Kennedy pathway for triacylglycerol (TAG) biosynthesis and, as such, elucidating its mode of regulation is critical to understand the fundamental aspects of carbon metabolism in oleaginous crops. In this study, purified Brassica napus Diacylglycerol Acyltransferase 1 (BnaDGAT1) in n-dodecyl-β-d-maltopyranoside micelles was lipidated to form mixed micelles and subjected to detailed biochemical analysis. The degree of mixed micelle fluidity appeared to influence Acyltransferase activity. BnaDGAT1 exhibited a sigmoidal response and eventual substrate inhibition with respect to increasing concentrations of oleoyl-CoA. Phosphatidate (PA) was identified as a feed-forward activator of BnaDGAT1, enabling the final enzyme in the Kennedy pathway to adjust to the incoming flow of carbon leading to TAG. In the presence of PA, the oleoyl-CoA saturation plot became more hyperbolic and desensitized to substrate inhibition indicating that PA facilitates the transition of the enzyme into the more active state. PA may also relieve possible autoinhibition of BnaDGAT1 brought about by the N-terminal regulatory domain, which was shown to interact with PA. Indeed, PA is a key effector modulating lipid homeostasis, in addition to its well recognized role in lipid signaling. BnaDGAT1 was also shown to be a substrate of the sucrose non-fermenting-1-related kinase 1 (SnRK1), which catalyzed phosphorylation of the enzyme and converted it to a less active form. Thus, this known regulator of carbon metabolism directly influences TAG biosynthesis.
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Purification and properties of recombinant Brassica napus Diacylglycerol Acyltransferase 1
FEBS Letters, 2015Co-Authors: Kristian Mark P. Caldo, Guanqun Chen, M. Joanne Lemieux, Michael S. Greer, Randall J. WeselakeAbstract:Diacylglycerol Acyltransferase 1 (DGAT1) catalyzes the final step in the acyl-CoA-dependent triacylglycerol biosynthesis. Although the first DGAT1 gene was identified many years ago and the encoded enzyme catalyzes a key step in lipid biosynthesis, no detailed structure–function information is available on the enzyme due to difficulties associated with its purification. This study describes the purification of recombinant Brassica napus DGAT1 (BnaC.DGAT1.a) in active form through solubilization in n-dodecyl-β-d-maltopyranoside, cobalt affinity chromatography, and size-exclusion chromatography. Different BnaC.DGAT1.a oligomers in detergent micelles were resolved during the size-exclusion process. BnaC.DGAT1.a was purified 126-fold over the solubilized fraction and exhibited a specific activity of 26 nmol TAG/min/mg protein. The purified enzyme exhibited substrate preference for α-linolenoyl-CoA > oleoyl-CoA = palmitoyl-CoA > linoleoyl-CoA > stearoyl-CoA.
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antisense suppression of type 1 Diacylglycerol Acyltransferase adversely affects plant development in brassica napus
Physiologia Plantarum, 2009Co-Authors: Yeeying Lock, Randall J. Weselake, Crystal L Snyder, Rodrigo M P Siloto, Saleh ShahAbstract:Diacylglycerol Acyltransferase (DGAT) catalyzes the acyl-coenzyme A (CoA) dependent acylation of sn-1,2-Diacylglycerol to form triacylglycerol in the terminal step of seed oil formation. Previous work has suggested that the level of DGAT activity may have a substantial effect on the flow of carbon into triacylglycerol, implying that the enzyme may represent a promising target for seed oil modification through biotechnological approaches. In the current study, Brassica napus DH12075 was transformed with an antisense type 1 DGAT construct, resulting in a reduction in DGAT1 gene expression, total DGAT activity and seed oil content. In addition, reduced seed yield and germination rates were observed along with severe developmental abnormalities. These data suggest that in addition to its critical role in seed oil formation, DGAT1 enzyme may also be important for normal seed development in B. napus, although the underlying mechanism(s) remain to be determined.
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Diacylglycerol Acyltransferase: A key mediator of plant triacylglycerol synthesis
Lipids, 2006Co-Authors: Shiu-cheung Lung, Randall J. WeselakeAbstract:Many plants deposit TAG in seeds and fruits as the major form of storage lipid. TAG production is of tremendous socioeconomic value in food, nutraceutical, and industrial applications, and thus numerous conventional and molecular genetic strategies have been explored in attempts to increase TAG content and modify the FA composition of plant seed oils. Much research has focused on the acyl-CoA-dependent reaction catalyzed by Diacylglycerol Acyltransferase (DGAT), which is an integral endoplasmic reticulum protein and has also been shown to be present in oil bodies and plastids. DGAT enzymes exhibit diverse biochemical properties among different plant species, many of which are summarized here. In addition to catalyzing a critical step in TAG biosynthesis, there is evidence that DGAT has roles in lipid metabolism associated with germination and leaf senescence. TAG can also be formed in plants via two different acyl-CoA-independent pathways, catalyzed by phospholipid: Diacylglycerol Acyltransferase and Diacylglycerol transacylase. The current understanding of the terminal step in TAG formation in plants and the development of molecular genetic approaches aimed at altering TAG yield and FA composition of TAG are discussed.
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Stability of Diacylglycerol Acyltransferase in dehydrated bovine muscle tissue
Analytical Biochemistry, 2003Co-Authors: Phillip W Huff, Fred J. Lozeman, E.chris Kazala, Cara Prozniak, Jochen Wegner, Jingti Deng, André Laroche, Jennifer L. Aalhus, Randall J. WeselakeAbstract:Meaningful estimates of Diacylglycerol Acyltransferase (EC 2.3.1.20) activity in different tissue samples require effective, unbiased methods of sample storage. Samples of the pars costalis diaphragmatis muscle (skirt muscle of the diaphragm) were obtained from 18- to 20-month-old cattle and assayed for microsomal protein content and Diacylglycerol Acyltransferase activity after having been stored under various conditions as dissected tissue or microsomes prepared from dissected tissue. There was relative enrichment of Diacylglycerol Acyltransferase specific activity (p
Ljerka Kunst - One of the best experts on this subject based on the ideXlab platform.
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identification of the wax ester synthase acyl coenzyme a Diacylglycerol Acyltransferase wsd1 required for stem wax ester biosynthesis in arabidopsis
Plant Physiology, 2008Co-Authors: Fengling Li, Xuemin Wu, David F Bird, Huanquan Zheng, Lacey Samuels, Reinhard Jetter, Ljerka KunstAbstract:Wax esters are neutral lipids composed of aliphatic alcohols and acids, with both moieties usually long-chain (C16 and C18) or very-long-chain (C20 and longer) carbon structures. They have diverse biological functions in bacteria, insects, mammals, and terrestrial plants and are also important substrates for a variety of industrial applications. In plants, wax esters are mostly found in the cuticles coating the primary shoot surfaces, but they also accumulate to high concentrations in the seed oils of a few plant species, including jojoba (Simmondsia chinensis), a desert shrub that is the major commercial source of these compounds. Here, we report the identification and characterization of WSD1, a member of the bifunctional wax ester synthase/Diacylglycerol Acyltransferase gene family, which plays a key role in wax ester synthesis in Arabidopsis (Arabidopsis thaliana) stems, as first evidenced by severely reduced wax ester levels of in the stem wax of wsd1 mutants. In vitro assays using protein extracts from Escherichia coli expressing WSD1 showed that this enzyme has a high level of wax synthase activity and approximately 10-fold lower level of Diacylglycerol Acyltransferase activity. Expression of the WSD1 gene in Saccharomyces cerevisiae resulted in the accumulation of wax esters, but not triacylglycerol, indicating that WSD1 predominantly functions as a wax synthase. Analyses of WSD1 expression revealed that this gene is transcribed in flowers, top parts of stems, and leaves. Fully functional yellow fluorescent protein-tagged WSD1 protein was localized to the endoplasmic reticulum, demonstrating that biosynthesis of wax esters, the final products of the alcohol-forming pathway, occurs in this subcellular compartment.
Saleh Shah - One of the best experts on this subject based on the ideXlab platform.
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antisense suppression of type 1 Diacylglycerol Acyltransferase adversely affects plant development in brassica napus
Physiologia Plantarum, 2009Co-Authors: Yeeying Lock, Randall J. Weselake, Crystal L Snyder, Rodrigo M P Siloto, Saleh ShahAbstract:Diacylglycerol Acyltransferase (DGAT) catalyzes the acyl-coenzyme A (CoA) dependent acylation of sn-1,2-Diacylglycerol to form triacylglycerol in the terminal step of seed oil formation. Previous work has suggested that the level of DGAT activity may have a substantial effect on the flow of carbon into triacylglycerol, implying that the enzyme may represent a promising target for seed oil modification through biotechnological approaches. In the current study, Brassica napus DH12075 was transformed with an antisense type 1 DGAT construct, resulting in a reduction in DGAT1 gene expression, total DGAT activity and seed oil content. In addition, reduced seed yield and germination rates were observed along with severe developmental abnormalities. These data suggest that in addition to its critical role in seed oil formation, DGAT1 enzyme may also be important for normal seed development in B. napus, although the underlying mechanism(s) remain to be determined.
Fengling Li - One of the best experts on this subject based on the ideXlab platform.
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identification of the wax ester synthase acyl coenzyme a Diacylglycerol Acyltransferase wsd1 required for stem wax ester biosynthesis in arabidopsis
Plant Physiology, 2008Co-Authors: Fengling Li, Xuemin Wu, David F Bird, Huanquan Zheng, Lacey Samuels, Reinhard Jetter, Ljerka KunstAbstract:Wax esters are neutral lipids composed of aliphatic alcohols and acids, with both moieties usually long-chain (C16 and C18) or very-long-chain (C20 and longer) carbon structures. They have diverse biological functions in bacteria, insects, mammals, and terrestrial plants and are also important substrates for a variety of industrial applications. In plants, wax esters are mostly found in the cuticles coating the primary shoot surfaces, but they also accumulate to high concentrations in the seed oils of a few plant species, including jojoba (Simmondsia chinensis), a desert shrub that is the major commercial source of these compounds. Here, we report the identification and characterization of WSD1, a member of the bifunctional wax ester synthase/Diacylglycerol Acyltransferase gene family, which plays a key role in wax ester synthesis in Arabidopsis (Arabidopsis thaliana) stems, as first evidenced by severely reduced wax ester levels of in the stem wax of wsd1 mutants. In vitro assays using protein extracts from Escherichia coli expressing WSD1 showed that this enzyme has a high level of wax synthase activity and approximately 10-fold lower level of Diacylglycerol Acyltransferase activity. Expression of the WSD1 gene in Saccharomyces cerevisiae resulted in the accumulation of wax esters, but not triacylglycerol, indicating that WSD1 predominantly functions as a wax synthase. Analyses of WSD1 expression revealed that this gene is transcribed in flowers, top parts of stems, and leaves. Fully functional yellow fluorescent protein-tagged WSD1 protein was localized to the endoplasmic reticulum, demonstrating that biosynthesis of wax esters, the final products of the alcohol-forming pathway, occurs in this subcellular compartment.
Yuki Nakamura - One of the best experts on this subject based on the ideXlab platform.
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functional study of Diacylglycerol Acyltransferase type 2 family in chlamydomonas reinhardtii
FEBS Letters, 2013Co-Authors: Chunhsien Hung, Ming Yang Ho, Kazue Kanehara, Yuki NakamuraAbstract:Algal triacylglycerol biosynthesis is of increasing interest for potential biodiesel production. A model microalga, Chlamydomonas, has multiple isoforms of Diacylglycerol Acyltransferase type 2 (DGTT) catalyzing the final step of triacylglycerol biosynthesis; however, the functions of the isoforms are poorly understood. Here, we performed heterologous complementation assay of Chlamydomonas DGTT1 to 4 in a yeast mutant defective in triacylglycerol biosynthesis. DGTT1, 2 and 3 but not 4 complemented the phenotype, including triacylglycerol levels. Interestingly, complementation by DGTT2 increased triacylglycerol content by 9-fold.