Figure 2.

SRD005825 inhibits binding of 9-cis-retinal by wild-type opsin. (A) Competition assay: purified human wild-type opsin was diluted to 0.12 mg/mL (3.0 μM) and incubated with 0, 20, 40, or 80 μM SRD005825 for 5 minutes followed by the addition of 2.0 μM 9-cis-retinal. Absorbance at 490 and 600 nm was measured every 2 minutes for 60 minutes to monitor the formation of rhodopsin. To eliminate background noise, the absorbance value at 600 nm was subtracted from the absorbance value at 490 nm. (B) Initial rates of binding: the initial rate of reaction was determined using a negative logarithm of the fraction of maximum rhodopsin formation (Log[A_max−A]/A_max, where A is adjusted absorbance at 490 nm).33,34 (C) Results of three separate competition assays. IC₅₀ values are estimated because complete competition was not achieved.