PHOTOAFFINITY-LABELING OF RETINOIC ACID-BINDING PROTEINS

Retinoid-binding proteins are essential mediators of vitamin A functio n in vertebrate organisms. They solubilize and stabilize retinoids, an d they direct the intercellular and intracellular trafficking, transpo rt, and metabolic function of vitamin A compounds in vision and in gro wth and development. Although many soluble retinoid-binding proteins a nd receptors have been purified and extensively characterized, relativ ely few membrane-associated enzymes and other proteins that interact w ith retinoids have been isolated and studied, due primarily to their i nherent instabilities during purification. In an effort to identify an d purify previously uncharacterized retinoid-binding proteins, it is s hown that radioactively labeled all-trans-retinoic acid can be used as a photoaffinity labeling reagent to specifically tag two known retino ic acid-binding proteins, cellular retinoic acid-binding protein and a lbumin, in complex mixtures of cytosolic proteins. Additionally, a num ber of other soluble and membrane-associated proteins that bind all-tr ans-[11,12-H-3] retinoic acid with high specificity are labeled utiliz ing the same photoaffinity techniques. Most of these labeled proteins have molecular weights that do not correspond to any known retinoid-bi nding proteins. Thus, photoaffinity labeling with all-trans-retinoic a cid and related photoactivatable retinoids is a method that should pro ve extremely useful in the identification and purification of novel so luble and membrane-associated retinoid-binding proteins from ocular an d nonocular tissues.