Specific inactivation of isomerohydrolase activity by 11-cis-retinoids

The endergonic trans-->cis isomerization of retinoids is an essential element in rhodopsin regeneration in vertebrates. All-trans-retinyl esters, which are generated by lecithin retinol acyltransferase (LRAT), are on the isomerization pathway. The critical isomerohydrolase activity, which catalyzes the trans-->cis isomerization/hydrolysis reaction of all-trans-retinyl esters, remains to be identified. It is demonstrated here that 11-cis-retinyl bromoacetate (cRBA) is a potent and specific inactivator of the bovine retinyl pigment epithelial (RPE) isomerohydrolase activity, with a measured K-I=0.19 muM and a pseudo-first-order rate of inactivation k(inh)=1.83x10(-3) s(-1). This demonstrates that the isomerization is indeed enzyme-mediated. This inactivator should facilitate the identification and study of isomerohydrolase, or at least an essential component of it. Labeling of crude RPE membranes with H-3-cRBA reveals the presence of several labeled bands that may be isomerohydrolase candidates. (C) 2003 Elsevier B.V. All rights reserved.