Binding analysis of amino-terminal and carboxyl-terminal regions of the 39-kDa protein to the low density lipoprotein receptor-related protein.
A 39-kDa protein binds to the low density lipoprotein receptor-related protein/alpha 2-macroglobulin receptor (LRP/alpha 2MR) and inhibits the binding of ligands to this receptor. We recently reported that inhibition of tissue-type plasminogen activator binding to LRP/alpha 2MR is mediated by both amino-terminal and carboxyl-terminal regions of the 39-kDa protein, whereas inhibition of alpha 2-macroglobulin-proteinase binding is mediated only by amino-terminal regions. In this report we show that amino-terminal and carboxyl-terminal regions of the 39-kDa protein bind specifically and with high affinity to LRP/alpha 2MR on rat hepatoma MH1C1 cells. Following binding, these amino-terminal and carboxyl-terminal regions of the 39-kDa protein are each rapidly endocytosed and degraded with kinetics identical to the full-length 39-kDa protein. Competition binding experiments with these constructs demonstrate that amino-terminal and carboxyl-terminal regions of the 39-kDa protein compete with one another for binding to LRP/alpha 2MR. A model is proposed in which amino-terminal and carboxyl-terminal regions of the 39-kDa protein bind to different sites on LRP/alpha 2MR in order to inhibit ligand binding.
Warshawsky
I
I
Edward Mallinckrodt Department of Pediatrics, Washington University School of Medicine, St. Louis, Missouri 63110.
Bu
G
G
Schwartz
A L
AL
D000818:Animals; D001665:Binding Sites; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D003001:Cloning, Molecular; D004705:Endocytosis; D005982:Glutathione Transferase / Q000378:metabolism; D006023:Glycoproteins / Q000378:metabolism*; D007700:Kinetics; D026563:LDL-Receptor Related Protein-Associated Protein; D008113:Liver Neoplasms; D008114:Liver Neoplasms, Experimental; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D008970:Molecular Weight; D010446:Peptide Fragments / Q000378:metabolism / Q000494:pharmacology; D011485:Protein Binding; D011869:Radioligand Assay; D051381:Rats; D011971:Receptors, Immunologic / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013379:Substrate Specificity; D010959:Tissue Plasminogen Activator / Q000378:metabolism; D014407:Tumor Cells, Cultured; D000511:alpha-Macroglobulins / Q000378:metabolism*
D002352:Carrier Proteins; D006023:Glycoproteins; D026563:LDL-Receptor Related Protein-Associated Protein; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D010446:Peptide Fragments; D011971:Receptors, Immunologic; D011993:Recombinant Fusion Proteins; D000511:alpha-Macroglobulins; D005982:Glutathione Transferase; D010959:Tissue Plasminogen Activator