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P. D. Kiser, D. T. Lodowski, M. R. Chance, K. Palczewski; Crystal Structure of the Visual Cycle Isomerase RPE65. Invest. Ophthalmol. Vis. Sci. 2009;50(13):1211.
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RPE65 is an iron-dependent, microsomal membrane protein that is exclusively expressed in the retinal pigmented epithelium and performs the critical trans-to-cis retinoid isomerization reaction of the visual cycle. Although much progress has been made in understanding RPE65 function we still do not fully understand the basic biochemical mechanism of retinoid isomerization. The structure of RPE65 may provide critical information regarding the active site architecture as well as provide a foundation for structure-based design of new RPE65 inhibitors.
RPE65 was used to produce diffraction-quality crystals. A complete diffraction dataset was collected to better than 2.15 angstrom resolution. The structure was solved using the single wavelength anomalous scattering technique and refined to an Rfree value of 23%.
RPE65 crystallizes with two monomers in the asymmetric unit. The protein adopts a seven-bladed beta-propeller fold. A mononuclear iron ion is found in the center of the beta propeller coordinated by four highly conserved histidine residues. The channel through which substrate travels to reach the active site is clear from the structure as is the surface of the protein that is responsible for membrane binding. The residues that make up the active site are all well resolved in the electron density maps.
The high resolution data used in this study will allow us to make proposals regarding the RPE65 enzymatic mechanism. Additionally, this structure may be useful for the design of new, highly specific and high affinity RPE65 inhibitors for the treatment of Stargardt disease and dry-type age-related macular degeneration (AMD).
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