IDENTIFICATION OF DOMAINS ON THE 39-KDA PROTEIN THAT INHIBIT THE BINDING OF LIGANDS TO THE LOW-DENSITY-LIPOPROTEIN RECEPTOR-RELATED PROTEIN

The low density lipoprotein receptor-related protein/alpha2-macroglobu lin receptor (LRP/alpha2MR) binds and internalizes several plasma prot eins including tissue-type plasminogen activator (t-PA) and alpha2-mac roglobulin-protease complexes (alpha2M). A 39-kDa protein that copuri fies with LRP/alpha2MR inhibits the binding and uptake of ligands by L RP/alpha2MR, including t-PA and alpha2M. To define domains on the 39- kDa protein which are essential for inhibition of t-PA and alpha2M bi nding to LRP/alpha2MR, we have generated bacterial expression construc ts encoding discrete regions of the 39-kDa protein as fusion proteins with glutathione S-transferase. Inhibition of t-PA and alpha2M bindin g to LRP/alpha2MR on rat hepatoma MH1C1 cells are shown to require ami no acid residues 18-24 and 100-107 on the 39-kDa protein. Inhibition o f t-PA but not alpha2M binding to LRP/alpha2MR is also mediated by re sidues 200-225 and 311-319. The same 39-kDa protein constructs that in hibit alpha2M and t-PA binding to MH1C1 cells are able to bind direct ly to purified LRP/alpha2MR immobilized on nitrocellulose. Thus, our s tudies demonstrate several specific regions on the 39-kDa protein whic h are required for the inhibition of t-PA and alpha2M binding to LRP/ alpha2MR.