Writing in Cell, Jin and colleagues report that they have identified the long sought-after isomerase enzyme responsible for the regeneration of the visual pigment rhodopsin in the retina after light exposure.

Light perception in vertebrates is mediated by rhodopsin, which is embedded in the disc membrane of rod cells. Rhodopsin is composed of a protein portion, opsin, which is a member of the G-protein-coupled receptor family, and a light-absorbing portion, 11-cis-retinaldehyde, which is a derivative of vitamin A.

Activation of rhodopsin starts with the absorption of light by 11-cis-retinaldehyde, which is then converted to all-trans-retinaldehyde and subsequently dissociates from opsin through a process known as photobleaching. Opsin itself is insensitive to light, and the regeneration of rhodopsin depends on a cascade of enzymatic reactions that converts all-trans-retinaldehyde back to 11-cis-retinaldehyde, completing the visual cycle. The key step in this cycle is the conversion of all-trans-retinyl palmitate into 11-cis-retinol, which is catalysed by an isomerase that has eluded researchers for more than two decades.

In this study, Jin et al. carried out an unbiased functional screen and discovered that RPE65 (retinal pigment epithelium-specific protein 65), which was thought to present the substrate to the elusive enzyme, was the isomerase itself. To confirm this finding, RPE65 was expressed in mammalian and insect cells alone or in combination with other enzymes of the visual cycle. RPE65 alone was sufficient to catalyse the conversion of all-trans-retinyl palmitate into all-cis-retinol, which indicates that RPE65 has intrinsic isomerase activity. This conclusion is also supported by in vitro isomerase assays using membrane extracts from cells expressing RPE65.

Mutations in the RPE65 gene are associated with Leber congenital amaurosis, an recessively inherited disease that is believed to cause up to 20% of all cases of childhood blindness. The researchers used site-directed mutagenesis to generate alleles of RPE65 associated with the disease, and found that cells expressing these mutants could not convert 11-trans-retinyl palmitate into 11-cis-retinol. The protein levels of wild-type and mutated RPE65 were similar in these cells, which indicates that lack of 11-cis-retinol production was due to reduced catalytic activity of RPE65 rather than a defect in protein expression or stability.

These findings have solved the long-standing mystery about the identity of the retinoid isomerase, and have significant implications for treating congenital blindness.