ANALYSIS OF LIGAND-BINDING TO THE ALPHA(2)-MACROGLOBULIN RECEPTOR LOW-DENSITY-LIPOPROTEIN RECEPTOR-RELATED PROTEIN - EVIDENCE THAT LIPOPROTEIN LIPASE AND THE CARBOXYL-TERMINAL DOMAIN OF THE RECEPTOR-ASSOCIATED PROTEIN BIND TO THE SAME SITE

The endocytic alpha(2)-macroglobulin receptor/low density lipoprotein receptor-related protein (alpha(2)MR/LRP) binds several classes of extracellular ligands at independent sites. In addition, alpha(2)MR/LRP can bind multiple copies of the 39-40-kDa receptor-associated protein (RAP). Both amino-terminal and carboxyl-terminal fragments of RAP exhibit affinity, and the fragments apparently bind to different sites on the receptor. RAP completely inhibits the binding of all presently known extracellular ligands, whereas several ligands such as alpha(2)-macroglobulin and tissue type plasminogen activator are poor inhibitors of RAP binding. Since RAP is largely an intracellular molecule that normally does not occupy alpha(2)MR/LRP at the cell surface, we hypothesized that an established extracellular ligand might bind to those sites on the receptor capable of binding the RAP fragments. We found complete cross competition between carboxyl-terminal RAP fragments and fragments of lipoprotein lipase containing the recently identified binding domain for alpha(2)MR/LRP (Nykjaer, A., Nielsen, M., Lookene, A., Meyer, N., Roigaard, H., Etzerodt, M., Beisiegel, U., Olivecrona, G., and Gliemann, J. (1994) J. Biol. Chem. 269, 31747-31755). Moreover, the lipoprotein lipase fragment completely inhibited the binding of several alpha(2)MR/LRP ligands in a pattern similar to that of carboxyl-terminal RAP fragments. On the other hand, the amino-terminal RAP fragment was a poor competitor of binding of the lipoprotein lipase fragment, whereas it competed effectively with pro-uPA for bind ing to the receptor. The results provide evidence that lipoprotein lipase binds to the site on alpha(2)MR/LRP also available for binding of the carboxyl terminal domain of RAP and suggest that pro-uPA may bind to or overlap the site available for the amino-terminal domain of RAP.