STRUCTURAL-ANALYSIS OF THE 6TH IMMUNOGLOBULIN-LIKE DOMAIN OF MOUSE NEURAL CELL-ADHESION MOLECULE L1 AND ITS INTERACTIONS WITH ALPHA-V-BETA-3, ALPHA-IIB-BETA-3, AND ALPHA-5-BETA-1 INTEGRINS

Previous experiments suggested that the human cell adhesion molecule L 1 interacts with different integrins via its sixth immunoglobulin-like domain in an RGD-dependent manner. Here we have described the express ion of this domain from early postnatal mouse brain, analyzed the stru cture of the recombinant protein by circular dichroism and fluorescenc e spectroscopy, and performed solid-phase binding studies to alpha v b eta 3, alpha IIb beta 3, and alpha 5 beta 1 integrins. The domain was found to have the expected P-sheet organization, which was lost in the presence of guanidine hydrochloride. The midpoint of the single-step transition occurred at 1.5 M guanidine hydrochloride. The sixth immuno globulin-like domain of mouse brain L1 contains two RGD motifs and was found to bind in a concentration-dependent and saturable way to alpha v beta 3, alpha IIb beta 3, and alpha 5 beta 1 integrins, suggesting specific interactions with these ligands. However, only the interactio n to alpha v beta 3 could be inhibited in a concentration-dependent ma nner by an RGD-containing peptide, and the IC50 was determined to be s imilar to 20 nM. Mutants of the domain, which lack either one or both of the RGD sites, demonstrated that the RGD site comprising residues 5 62-564 is involved in the interaction to alpha v beta 3, Our findings indicate an RGD-independent mechanism for the interactions to alpha II b beta 3 and alpha 5 beta 1, as no involvement of any RGD motif could be demonstrated.