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S. Bussières, R. Breton, C. Salesse; Membrane Binding of a Transmembrane Domain–Deleted Form of Lecithin Retinol Acyltransferase . Invest. Ophthalmol. Vis. Sci. 2006;47(13):2019.
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Lecithin retinol acyltransferase is a 230 amino acids transmembrane protein which catalyzes the esterification of all–trans–retinol into all–trans–retinyl ester, an essential reaction in the vertebrate visual cycle. The present study was performed to determine the interaction of the transmembrane domain deleted form of LRAT (tLRAT) with membranes and to determine its secondary structure.
The truncated form of LRAT (tLRAT) which extends from residues 31 to 196 has been produced as previously described. The deleted domains did not contain residues known to be required for catalysis. tLRAT has been purified using affinity chromatography. Polarization–modulated infrared reflection absorption spectroscopy (PM–IRRAS), surface–pressure adsorption isotherms and Brewster angle microscopy (BAM) have been used to measure tLRAT adsorption onto phospholipid monolayers at the air–water interface, to probe its secondary structure and lipid hydrolysis as well as to observe film structure.
tLRAT was found to strongly interact with lipid monolayers. The analysis of the PM–IRRAS spectra indicated that tLRAT contains a major proportion of alpha–helical structure when bound to lipid monolayers whereas it mainly contains a mixture of alpha–helical and beta structures in solution. PM–IRRAS also allowed to monitor lipid hydrolysis by LRAT. Upon adsorption, tLRAT forms a lace–like structure in monolayers as observed by BAM.
tLRAT interacts with membranes and then hydrolyzes membrane lipids. It changes conformation upon membrane binding and forms a homogeneously organized film.
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