ALL-TRANS RETINOL, VITAMIN-D AND OTHER HYDROPHOBIC COMPOUNDS BIND IN THE AXIAL PORE OF THE 5-STRANDED COILED-COIL DOMAIN OF CARTILAGE OLIGOMERIC MATRIX PROTEIN

The potential storage and delivery function of cartilage oligomeric ma trix protein (COMP) for cell signaling molecules was explored by bindi ng hydrophobic compounds to the recombinant five-stranded coiled-coil domain of COMP. Complex formation with benzene, cyclohexane, vitamin D -3 and elaidic acid was demonstrated through increases in denaturation temperatures of 2-10 degrees C, For all-trans retinol and all-trans r etinoic acid, an equilibrium dissociation constant K-D = 0.6 mu M was evaluated by fluorescence titration. Binding of benzene and all-trans retinol into the hydrophobic axial pore of the COMP coiled-coil domain was proven by the X-ray crystal structures of the corresponding compl exes at 0.25 and 0.27 nm resolution, respectively. Benzene binds with its plane perpendicular to the pore axis. The binding site is betvveen the two internal rings formed by Leu37 and Thr40 pointing into the po re of the COMP coiled-coil domain. The retinol beta-ionone ring is pos itioned in a hydrophobic environment near Thr40, and the 1.1 nm long i soprene tail follows a completely hydrophobic region of the pore. Its terminal hydroxyl group complexes with a ring of the five side chains of Gln54, A mutant in which Gln54 is replaced by Ile binds all-trans r etinol with affinity similar to the wild-type, demonstrating that hydr ophobic interactions are predominant.