Acyl-acyl carrier protein thioesterase activity from sunflower ( Helianthus annuus L.) seeds

Martı́nez-Force, Enrique; Cantisán, Sara; Serrano-Vega, María J.; Garcés, Rafael

doi:10.1007/s004250000332

Cited by 27 publications

(19 citation statements)

References 25 publications

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“…This comparison demonstrated that the protein encoded by HaFatA1 is highly homologous to FatA thioesterases, and more specifically with those of the ''campanulids'' group like Carthamus tinctorius (Knutzon et al 1992) and Coriandrum sativum (Do¨rmann et al 1994). Significantly, HaFatA1 was clearly different from the FatB thioesterases, confirming the presence of a FatA type thioesterase in developing sunflower seeds as previously proposed by Martı´nez-Force et al (2000).…”

Section: Resultssupporting

confidence: 85%

“…The soluble and membrane-bound protein fractions from E. coli cells expressing FatA1 thioesterase were collected and assayed for thioesterase activity using 18:1-ACP, the preferred substrate for sunflower FatA thioesterase (Martı´nez-Force et al 2000). The acyl-ACP thioesterase activity was approximately 20,000 fold higher in cells containing pQEFATA than in control cells.…”

Section: Expression Of Sunflower Fata1 Thioesterase In Escherichia Colimentioning

confidence: 99%

“…Seeds were peeled and ground in extraction buffer containing 50 mM tris-HCl (pH 8.0) and 5 mM dithiothreitol (DTT), 1 g of tissue per 10 ml of buffer (Martı´nez-Force et al 2000). The protein concentration was measured using a Protein Assay Kit (Bio-Rad, Hercules, CA, USA) according to the manufacturer's instructions and using BSA as a standard.…”

Section: Acyl-acp Thioesterase Assaysmentioning

confidence: 99%

“…During sunflower seed development, both FatA and FatB thioesterases have been shown to be expressed through competence assays and by PCR amplification of thioesterase mRNA (Martı´nez-Force et al 2000). Here, we report the cloning, expression, purification and biochemical characterization of a FatA type thioesterase from developing sunflower seeds, demonstrating the relative importance of this thioesterase in the composition of sunflower oil.…”

Section: Introductionmentioning

confidence: 98%

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Cloning, characterization and structural model of a FatA-type thioesterase from sunflower seeds (Helianthus annuus L.)

Serrano-Vega

Garcés²,

Martı́nez-Force³

2005

Planta

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The substrate specificity of acyl-acyl carrier protein (ACP) thioesterases (EC 3.1.2.14) determines the fatty acids available for the biosynthesis of storage and membrane lipids in seeds. In order to determine the mechanisms involved in the biosynthesis of fatty acids in sunflower seeds (Helianthus annuus L.), we isolated, cloned and sequenced a cDNA clone of acyl-ACP thioesterase from developing sunflower seeds, HaFatA1. Through the heterologous expression of HaFatA1 in Escherichia coli we have purified and characterized this enzyme, showing that sunflower HaFatA1 cDNA encodes a functional thioesterase with preference for monounsaturated acyl-ACPs. The HaFatA1 thioesterase was most efficient (kcat/K(m)) in catalyzing oleoyl-ACP, both in vivo and in vitro. By comparing this sequence with those obtained from public databases, we constructed a phylogenetic tree that included FatA and FatB thioesterases, as well as related prokaryotic proteins. The phylogenetic relationships support the endosymbiotic theory of the origin of eukaryotic cells and the suggestion that eubacteria from the delta-subdivision were the guest cells in the symbiosis with archaea. These prokaryotic proteins are more homologous to plant FatB, suggesting that the ancient thioesterases were more similar to FatB. Finally, using the available structure prediction methods, a 3D model of plant acyl-ACP thioesterases is proposed that reflects the combined data from direct mutagenesis and chimera studies. In addition, the model was tested by mutating the residues proposed to interact with the ACP protein in the FatA thioesterase by site-directed mutagenesis. The results indicate that this region is involved in the stabilization of the substrate at the active site.

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Section: Resultssupporting

confidence: 85%

Section: Expression Of Sunflower Fata1 Thioesterase In Escherichia Colimentioning

confidence: 99%

Section: Acyl-acp Thioesterase Assaysmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 98%

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Cloning, characterization and structural model of a FatA-type thioesterase from sunflower seeds (Helianthus annuus L.)

Serrano-Vega

Garcés²,

Martı́nez-Force³

2005

Planta

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“…WoLF PSORT was used to identify the signal peptide and the cleavage site where the mature protein begins, a program that predicts protein subcellular localization (Horton et al, 2007), with the information available for acyl-ACP thioesterases from Arabidopsis thaliana (Dörmann et al, 1995), Helianthus annuus (Martínez-Force et., al, 2000;Serrano-Vega et al, 2005) Gossypium hirsutum (Huynh et al, 2002), Diploknema butyracea (Jha et al, 2006), Madhuca longifolia (Ghosh and Sen, 2007), Ricinus communis (Sánchez-García et al, 2010) and Macadamia tetraphylla (Moreno-Pérez et al, 2011). Val-55 was the best candidate to represent the N-terminal amino acid of the mature CsFatA protein, corresponding to a signal peptide of 58 amino acid residues (Fig.…”

Section: Isolation and Sequence Analysis Of Acyl-acp Thioesterases Frmentioning

confidence: 99%

Acyl-ACP thioesterases from Camelina sativa: Cloning, enzymatic characterization and implication in seed oil fatty acid composition

et al. 2014

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Acyl-acyl carrier protein (ACP) thioesterases are intraplastidial enzymes that terminate de novo fatty acid biosynthesis in the plastids of higher plants by hydrolyzing the thioester bond between ACP and the fatty acid synthesized. Free fatty acids are then esterified with coenzyme A prior to being incorporated into glycerolipids. Acyl-ACP thioesterases are classified into two families, FatA and FatB, which differ in their amino acid sequence and substrate specificity. Here, the FatA and FatB thioesterases from Camelina sativa seeds, a crop of interest in plant biotechnology, were cloned, sequenced and characterized. The mature proteins encoded by these genes were characterized biochemically after they were heterologously expressed in Escherichia coli and purified. Camelina sativa contained three different alleles of both the FatA and FatB genes. These genes were expressed most strongly in expanding tissues in which lipids are very actively synthesized, such as developing seed endosperm. The CsFatA enzyme displayed high catalytic efficiency on oleoyl-ACP and CsFatB acted efficiently on palmitoyl-ACP. The contribution of these two enzymes to the synthesis of Camelina sativa oil was discussed in the light of these results.

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Lipid Metabolism

Schmid

2004

Handbook of Plant Biotechnology

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Lipids are an immensely diverse group of compounds, technically including all biomolecules that dissolve more readily in non‐polar solvents than in water. This review will emphasisze two prominent groups of lipids: the acyl lipids, whose hydrophobic units are derived from fatty acids, and the isoprenoids, made up of five‐carbon isopentene units. Acyl lipids, in the form of the triacylglycerols stored as energy reserves, underpin the vegetable oil industry. In addition, acyl lipids that dominate the bilayers of plant membranes, are indispensable for the protective layers that prevent plant desiccation and pathogen attack, and include several signalling molecules. Isoprenoids are even more diverse than acyl lipids, with more than 25,000known. Two prominent groups are the sterols, which are important membrane constituents, and the carotenoids, which collect light for photosynthesis, protect plants from excessive light, attract pollinators and fruit dispersal agents, and provide vitamin A precursors for consumers. Biotechnology has provided both a tool for understanding plant lipid metabolism, and a means of altering quality and quantity of these constituents to better serve the needs of industry and human nutrition better.

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Acyl-acyl carrier protein thioesterase activity from sunflower ( Helianthus annuus L.) seeds

Cited by 27 publications

References 25 publications

Cloning, characterization and structural model of a FatA-type thioesterase from sunflower seeds (Helianthus annuus L.)

Cloning, characterization and structural model of a FatA-type thioesterase from sunflower seeds (Helianthus annuus L.)

Acyl-ACP thioesterases from Camelina sativa: Cloning, enzymatic characterization and implication in seed oil fatty acid composition

Lipid Metabolism

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