Abstract

Vitamin A movement between rod outer segment (ROS) membranes, interphotoreceptor retinoid-binding protein (IRBP), and liposomes was examined by two different methods. Equilibrium exchange of all-trans-retinol was followed by assessing the transfer of [3H]retinol from liposomes to ROS membranes as compared to a nontransferable marker, [14C]triolein. In the absence of IRBP, a rapid, spontaneous transfer of [3H] retinol to the ROS membranes occurred. In the presence of 2 microM IRBP, retinol transfer decreased by approximately one-half, whereas a similar concentration of bovine serum albumin had no effect on this spontaneous transfer. Kinetics of retinol transfer between single unilamellar vesicles were determined by the method of fluorescence energy transfer. The first order rate constant for this transfer was 0.85 s-1 at 22 degrees C at either pH 7.4 or pH 2.8. This rate was not affected by varying the concentration of acceptor vesicles 50-fold or by varying their concentration 10-fold at a constant ratio of donor-to-acceptor vesicles. The presence of IRBP as an additional acceptor did not change the rate. The transfer was temperature-dependent with an activation energy of 7.8 kcal/mol. The transfer rate appeared to be an increasing exponential function of ionic strength since high concentrations of NaCl decreased the transfer rate significantly. The transfer rate of retinol from IRBP to single unilamellar vesicles also followed first order kinetics with a rate constant of 0.11 s-1 at 22 degrees C, which was approximately 8 times slower than that of transfer between vesicles. We conclude that the transfer of all-trans-retinol between liposomes and membranes can be accomplished rapidly via the aqueous phase, and that IRBP retards rather than facilitates this transfer process.

Highlights

  • From the ‘iProrrrarn in Neuroscience. the GCullen Eve Institute, and the TDepartrnentof Medicine, BaylorCollege of Medicine, I ”

  • Because of its unique localization, trans-retinol wasfollowed by assessing the transferof it has been repeatedly suggested [1,2,3,4] that IRBP serves to i3H1retinolfrom liposomes to ROS membranes as com- transport retinoids between outer segments of the photorepared to a nontransferable mark[e1r4, C]triolein.In the ceptor cells and the retinal pigment epithelium, i.e. to transabsence of IRBP, a rapid, spontaneous transfeofr [‘HI portall-trans-retinol released from outer segments to the retinol to the ROS membranes occurred

  • The transfer rate of rate was a function of both hydrophobicity and hydrophilicity retinol from IRBP to single unilamellar vesicles of the transferred molecule, i.e. it increased both with defollowed first order kinetics with a rate constant of creased chain length and increased polarity of the molecule

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Summary

THEJOURNAOFLBIOLOGICAL CHEMISTRY

0 1989 by The American Society for Biochemistry and Molecular Biology, Inc. Vol 264,No 2, Isaue of January 15,pp. 928-935,1989 Printed in U.S.A. 0 1989 by The American Society for Biochemistry and Molecular Biology, Inc. Vol 264,No 2, Isaue of January 15,pp. 928-935,1989 Printed in U.S.A. Mechanism of Vitamin A Movement between Rod Outer Segments, Interphotoreceptor Retinoid-binding Protein, anLdiposomes*. The GCullen Eve Institute, and the TDepartrnentof Medicine, BaylorCollege of Medicine, I ” From the ‘iProrrrarn in Neuroscience. the GCullen Eve Institute, and the TDepartrnentof Medicine, BaylorCollege of Medicine, I ”

Vitamin A movement betweenrodoutersegment
EXPERIMENTAL PROCEDURES
Equilibrium Exchange Experiments
Fraction number
Acceptor vesicle tn mM
DISCUSSION
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