Abstract
Abstract: :
Purpose: RPE65 is a major membrane protein of the retinal pigment epithelium (1,2). While knockout studies in mice suggest an important role for this protein in the operation of the visual cycle in vivo (3), a biochemical function for this protein remains obscure. This study addresses this issue. Methods: In the current studies, RPE65 is specifically and covalently labeled by the retinyl ester analog (3R)- 3-[boc-lys(biotinyl)-O]-all trans-retinol chloroacetate 1 at low µM concentrations to explore the retinoid binding capacity of RPE65. Results: The affinity labeling of RPE65 by 1 is blocked by added retinoids, including analog 2. The stoichiometry of labeling is two, and two cysteine residues (C231 and C448) of RPE65 are specifically labeled by 1. The known homology of RPE65 to ß-carotene-15, 15'-dioxygenase may be relevant here, because a ß-carotene binding site contains the elements of two retinoid binding sites. Conclusions: These studies demonstrate that RPE65 is a retinoid binding protein. Absent any demonstrated enzymatic activity for RPE65, a plausible function for this protein may be to act as a retinyl ester binding protein. Retinyl ester binding proteins would mobilize these highly hydrophobic retinoids, making them available for further processing in the visual cycle. 1. Bavik, C. O., Busch, C., Eriksson, U. (1992) J. Biol. Chem. 267, 23035-23042. 2. Hamel, C. P., Tsilou, E., Pfeffer, B. A., Hooks, J. J., Detrick, B., Redmond, T. M. (1993) J. Biol. Chem. 268, 15751-15757. 3. Redmond, T. M., Yu, S., Lee, E., Bok, D., Hamasaki, D., Chen, N., Goletz, P., Ma, J.-X., Crouch, R., Pfeifer, K. (1998) Nature Genet. 20, 344-351.
Keywords: retinoids/retinoid binding proteins • retinal pigment epithelium