December 2002
Volume 43, Issue 13
Free
ARVO Annual Meeting Abstract  |   December 2002
Interactions of CRALBP With Other Visual Cycle Proteins
Author Affiliations & Notes
  • SK Bhattacharya
    Ophthalmic Research Cole Eye Institute Cleveland Clinic Foundation Cleveland OH
  • Z Wu
    Ophthalmic Research Cole Eye Institute Cleveland Clinic Foundation Cleveland OH
  • M Miyagi
    Ophthalmic Research Cole Eye Institute Cleveland Clinic Foundation Cleveland OH
  • KA West
    Ophthalmic Research Cole Eye Institute Cleveland Clinic Foundation Cleveland OH
  • Z Jin
    Ophthalmic Research Cole Eye Institute Cleveland Clinic Foundation Cleveland OH
  • M Nawrot
    Ophthalmology University of Washington Seattle WA
  • JC Saari
    Ophthalmology University of Washington Seattle WA
  • JW Crabb
    Ophthalmic Research Cole Eye Institute Cleveland Clinic Foundation Cleveland OH
  • Footnotes
    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
Investigative Ophthalmology & Visual Science December 2002, Vol.43, 4567. doi:
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    • Get Citation

      SK Bhattacharya, Z Wu, M Miyagi, KA West, Z Jin, M Nawrot, JC Saari, JW Crabb; Interactions of CRALBP With Other Visual Cycle Proteins . Invest. Ophthalmol. Vis. Sci. 2002;43(13):4567.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Abstract: : 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 
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