Ligand specificity in the CRAL-TRIO protein family
ABSTRACT Intracellular trafficking of hydrophobic ligands is often mediated by specific binding proteins. The CRAL-TRIO motif is common to several lipid binding proteins including the cellular retinaldehyde binding protein (CRALBP), the alpha-tocopherol transfer protein (alpha-TTP), yeast phosphatidylinositol transfer protein (Sec14p), and supernatant protein factor (SPF). To examine the ligand specificity of these proteins, we measured their affinity toward a variety of hydrophobic ligands using a competitive [(3)H]-RRR-alpha-tocopherol binding assay. Alpha-TTP preferentially bound RRR-alpha-tocopherol over all other tocols assayed, exhibiting a K(d) of 25 nM. Binding affinities of other tocols for alphaTTP closely paralleled their ability to inhibit in vitro intermembrane transfer and their potency in biological assays. All other homologous proteins studied bound alpha-tocopherol but with pronouncedly weaker (> 10-fold) affinities than alpha-TTP. Sec14p demonstrated a K(d) of 373 nM for alpha-tocopherol, similar to that for its native ligand, phosphatidylinositol (381 nM). Human SPF had the highest affinity for phosphatidylinositol (216 nM) and gamma-tocopherol (268 nM) and significantly weaker affinity for alpha-tocopherol (K(d) 615 nM). SPF bound [(3)H]-squalene more weakly (879 nM) than the other ligands. Our data suggest that of all known CRAL-TRIO proteins, only alphaTTP is likely to serve as the physiological mediator of alpha-tocopherol's biological activity. Further, ligand promiscuity observed within this family suggests that caution should be exercised when suggesting protein function(s) from measurements utilizing a single ligand.
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ABSTRACT: An asymmetric synthetic route for 1-iodofarnesane, a key intermediate for tocopherol side chain synthesis, starting from (+)-(R)-citronellal was developed. 1-Iodofamesane was prepared through eight steps in about 50% overall yield, and asymmetric transfer hydrogenation of the enal with a chiral organocatalyst was conducted as a stereoinduction step. To measure the stereoinduction level and optical purity of the product, a convenient analytical method was developed in which a phenylcarbamate derivative of the C-15 alcohol was found to be suitable to give proper polarity and UV-activity for chiral UV-HPLC analysis.Tetrahedron Letters 10/2014; 55(43):5895–5897. DOI:10.1016/j.tetlet.2014.08.085 · 2.39 Impact Factor
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ABSTRACT: We review our recent work on protein–ligand interactions in vitamin transporters of the Sec-14-like protein. Our studies focused on the cellular-retinaldehyde binding protein (CRALBP) and the α-tocopherol transfer protein (α-TTP). CRALBP is responsible for mobilisation and photo-protection of short-chain cis-retinoids in the dim-light visual cycle or rod photoreceptors. α-TTP is a key protein responsible for selection and retention of RRR-α-tocopherol, the most active isoform of vitamin E in superior animals. Our simulation studies evidence how subtle chemical variations in the substrate can lead to significant distortion in the structure of the complex, and how these changes can either lead to new protein function, or be used to model engineered protein variants with tailored properties. Finally, we show how integration of computational and experimental results can contribute in synergy to the understanding of fundamental processes at the biomolecular scale.CHIMIA International Journal for Chemistry 09/2014; 68(9). DOI:10.2533/chimia.2014.615 · 1.09 Impact Factor
Chapter: Vitamin E[Show abstract] [Hide abstract]
ABSTRACT: Tocochromanols are a small group of natural products synthesized exclusively by photosynthetic organisms and include the tocopherols and tocotrienols that are vitamin E, an essential lipid-soluble antioxidant in the human diet. The major source of vitamin E in the human diet is plant-derived products, which can vary by orders of magnitude in tocochromanol content and composition and hence vitamin E activity. In the past decade, tremendous progress has been made in our understanding of the molecular genetics of tocochromanol synthesis in plants and cyanobacteria, and all the biosynthetic genes of the core pathway have now been cloned and studied in detail. Significant progress has been made in engineering tocochromanol content and composition in plant tissues. Our understanding of tocochromanol function(s) during the plant life cycle has been advanced by the isolation and study of pathway mutants that accumulate specific intermediates or completely eliminate tocochromanols from the organism. Tocochromanols are absolutely essential for limiting lipid oxidation during seed desiccation, storage and germination, and the severe fitness impact of tocochromanol deficiency at this stage of the plant life cycle makes it obvious why tocochromanol synthesis has been conserved in all seed-bearing plants during evolution. However, the functions of tocochromanols in mature plant tissues are surprisingly more limited than had long been assumed, especially in regard with plant stress.Biosynthesis of vitamins in plants, Edited by Elsevier, advances in botanical research, 01/2011: pages 179-227; , ISBN: 978-0-12-385853-5