title: Real-time analyses of retinol transport by the membrane receptor of plasma retinol binding protein.
abstract: Vitamin A is essential for vision and the growth/differentiation of almost all human organs. Plasma retinol binding protein (RBP) is the principle and specific carrier of vitamin A in the blood. Here we describe an optimized technique to produce and purify holo-RBP and two real-time monitoring techniques to study the transport of vitamin A by the high-affinity RBP receptor STRA6. The first technique makes it possible to produce a large quantity of high quality holo-RBP (100%-loaded with retinol) for vitamin A transport assays. High quality RBP is essential for functional assays because misfolded RBP releases vitamin A readily and bacterial contamination in RBP preparation can cause artifacts. Real-time monitoring techniques like electrophysiology have made critical contributions to the studies of membrane transport. The RBP receptor-mediated retinol transport has not been analyzed in real time until recently. The second technique described here is the real-time analysis of STRA6-catalyzed retinol release or loading. The third technique is real-time analysis of STRA6-catalyzed retinol transport from holo-RBP to cellular retinol binding protein I (CRBP-I). These techniques provide high sensitivity and resolution in revealing RBP receptor's vitamin A uptake mechanism.
authors: Department of Physiology, Jules Stein Eye Institute and Howard Hughes Medical Institute, University of California, Los Angeles, USA.
authors: Riki
authors: R
authors: Kawaguchi
authors: Ming
authors: M
authors: Zhong
authors: Hui
authors: H
authors: Sun
mesh_terms: D002851:Chromatography, High Pressure Liquid; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D054839:Retinol-Binding Proteins, Plasma / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D014801:Vitamin A / Q000378:metabolism*
chemical_list: D008565:Membrane Proteins; D011994:Recombinant Proteins; D054839:Retinol-Binding Proteins, Plasma; C519685:STRA6 protein, human; D014801:Vitamin A