ACTIVE-SITE-DIRECTED INACTIVATION OF CONSTITUTIVELY ACTIVE MUTANTS OFRHODOPSIN

Recently, mutations of the active site Lys(296) residue in rhodopsin ( Lys(296) --> Glu and Lys(296) --> Met) have been found as the cause of disease in some patients with autosomal dominant retinitis pigmentosa . In vitro, these mutations result in constitutive activation of the p rotein. In an effort to develop a potential therapeutic agent for trea tment of the disease, we have examined various amine derivatives of 11 -cis- and 9-cis-retinal for ability to irreversibly inactivate a relat ed constitutively active mutant, K296G. Three amines were prepared by reductive amination of retinal: 11-cis-retinylpropylamine, 11-cis-reti nylamine, and 9-cis-retinylamine. All three compounds inactivated K296 G, and the inactivation could not be reversed upon exposure to light. None of the compounds inactivated the wild-type protein. Although the amines were not effective on the naturally occurring retinitis pigment osa mutants, presumably because of unfavorable steric interactions wit h the bulky Glu and Met side chains at position 296, the success with K296G makes it highly encouraging that this approach will evolve relat ed compounds that are capable of inactivating the naturally occurring mutants as well.