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Invest Ophthalmol Vis Sci 2002;43: E-Abstract 4567.
© 2002 ARVO

Interactions of CRALBP With Other Visual Cycle Proteins

SK Bhattacharya1, Z Wu1, M Miyagi1, KA West1, Z Jin1, M Nawrot2, JC Saari2 and JW Crabb1

1 Ophthalmic Research Cole Eye Institute Cleveland Clinic Foundation Cleveland OH
2 Ophthalmology University of Washington Seattle WA

Commercial Relationships:  S.K. Bhattacharya, None; Z. Wu, None; M. Miyagi, None; K.A. West, None; Z. Jin, None; M. Nawrot, None; J.C. Saari, None; J.W. Crabb, None.
Grant Identification: NIH Grants EY06603, EY01730, EY02317, a Research Center Grant from The foundation Fighting Blindness


Purpose:To demonstrate an interaction between cellular retinaldehyde-binding protein (CRALBP) and 11-cis-retinol-dehydrogenase (RDH5) and to identify other visual cycle protein interactions. The visual cycle is the enzymatic pathway by which all-trans-retinal from photoreceptor bleaching is isomerized to 11-cis-retinal in the retinal pigment epithelium (RPE) for visual pigment regeneration.
Methods:Recombinant CRALBP and RDH5 were produced and purified to apparent homogeneity. Kinetic parameters for RDH5 catalyzed reactions with retinoid substrates were determined in the presence and absence of CRALBP. Protein-protein interactions were sought in bovine RPE microsomes by reciprocal immunoprecipitations and other affinity isolation methods. Proteins were identified by Western analyses, MALDI TOF MS and LC MS/MS.
Results:Wildtype CRALBP enhances the affinity of RDH5 for 9-cis- and 11-cis-retinoids by 2-3 fold over free retinoid. Mutant M225K rCRALBP, which lacks ligand binding capability, has no effect on RDH5 enzyme kinetics. CRALBP, RDH5, RPE65 and RGR opsin co-precipitate from RPE microsomes with anti-CRALBP antibodies. Similar results were obtained with anti-RDH5 and anti-RPE65 antibodies.
Conclusion:A stable, functional interaction has been demonstrated between CRALBP and RDH5. Visual cycle proteins CRALBP, RDH5, RPE65 and RGR opsin appear to interact in a protein complex. Further proteomic and affinity isolation approaches are directed toward identifying other possible RPE components of a visual cycle protein complex.
CR: N. Supported in part by NIH grants EY06603, EY01730, EY02317, a Research Center Grant from The Foundation Fighting Blindness, and funds from the Cleveland Clinic Foundation and Research to Prevent Blindness, Inc.

Keywords: 527 protein structure/function • 571 retinoids/retinoid binding proteins • 567 retinal pigment epithelium

 © 2002, The Association for Research in Vision and Ophthalmology, Inc., all rights reserved. For permission to reproduce any part of this abstract, contact the ARVO Office at arvo{at}