Interconversion of red opsin isoforms by the cyclophilin-related chaperone protein Ran-binding protein 2

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Abstract
Ran-binding protein 2 (RanBP2) (type II) is a retinal cyclophilin-related protein that binds Ran-GTPase. Type I cyclophilin is a shorter, alternatively spliced isoform of RanBP2. Recently, we showed that the Ran-binding domain 4 (RBD4)/cyclophilin (CY) supradomain of RanBP2 acts both in vitro and in vivo as a specific chaperone for bovine red/green opsin (R/G opsin). R/G opsin undergoes a stable modification of its electrophoretic mobility upon binding to RanBP2. This modification is likely due to cis-trans isomerization of one or more proline residues in the opsin protein. Here, we show that expression of human red opsin in Escherichia coli and COS cells results in the production of still a third electrophoretic variant of this protein. This variant was converted to the RBD4 binding-competent form of opsin through direct interaction with RBD4/CY, both in vivo and in vitro. We suggest that these distinct opsin species may represent kinetically or thermodynamically trapped prolyl conformers that can be interconverted by concerted action of the RBD4 and CY domains of RanBP2. We also show that the C-terminal half of RBD4 is the binding domain for bovine R/G opsin and that coexpression of human red opsin with type I cyclophilin in vivo enhances the production of functional visual pigment. These observations imply that prolyl isomerization may have importance beyond its role in protein folding, possibly as a molecular switch modulated by cyclophilin for the loading of opsin onto RanBP2 during visual pigment processing in cones.