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Proc Natl Acad Sci U S A. Apr 12, 1994; 91(8): 3161–3165.
PMCID: PMC43535

Functional domains of the receptor-associated protein (RAP).

Abstract

The receptor-associated protein (RAP) specifically associates with gp330 and the low density lipoprotein (LDL) receptor-related protein (LRP), the two newest members of the LDL receptor gene family. Results obtained by ligand blotting, affinity chromatography, and density-gradient sedimentation demonstrate that RAP binds to both receptors with high affinity and that the binding is Ca2+ dependent. RAP also binds heparin and is identical to a mouse heparin binding protein (HBP-44) identified in a teratocarcinoma cell line (F9). While biochemical studies have shown that RAP is present on the cell surface and is an effective inhibitor of ligand binding to gp330 and LRP, immunocytochemical findings indicate that RAP is most abundant in the endoplasmic reticulum lumen and may function in receptor folding and/or trafficking. To facilitate the characterization of RAP's function(s) we have mapped its gp330 and heparin binding sites by performing direct binding studies on fusion proteins representing overlapping domains of RAP. gp330 was found to bind to two separate sites on RAP--i.e., between amino acids 85-148 and 178-248. Binding studies with radiolabeled heparin indicate that the heparin binding site is between amino acids 261 and 323, which is consistent with our previously proposed site (residues 287-306) based on the amphipathic nature of the C terminus of RAP. These data demonstrate that the gp330 and heparin binding sites and the Heymann nephritis pathogenic epitope (amino acids 1-86) demonstrated earlier are represented by distinct domains of the RAP polypeptide.

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Selected References

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  • Pietromonaco S, Kerjaschki D, Binder S, Ullrich R, Farquhar MG. Molecular cloning of a cDNA encoding a major pathogenic domain of the Heymann nephritis antigen gp330. Proc Natl Acad Sci U S A. 1990 Mar;87(5):1811–1815. [PMC free article] [PubMed]
  • Strickland DK, Ashcom JD, Williams S, Battey F, Behre E, McTigue K, Battey JF, Argraves WS. Primary structure of alpha 2-macroglobulin receptor-associated protein. Human homologue of a Heymann nephritis antigen. J Biol Chem. 1991 Jul 15;266(20):13364–13369. [PubMed]
  • Kanalas JJ, Makker SP. Analysis of a 45-kDa protein that binds to the Heymann nephritis autoantigen GP330. J Biol Chem. 1993 Apr 15;268(11):8188–8192. [PubMed]
  • Lundstrom M, Orlando RA, Saedi MS, Woodward L, Kurihara H, Farquhar MG. Immunocytochemical and biochemical characterization of the Heymann nephritis antigenic complex in rat L2 yolk sac cells. Am J Pathol. 1993 Nov;143(5):1423–1435. [PMC free article] [PubMed]
  • Orlando RA, Kerjaschki D, Kurihara H, Biemesderfer D, Farquhar MG. gp330 associates with a 44-kDa protein in the rat kidney to form the Heymann nephritis antigenic complex. Proc Natl Acad Sci U S A. 1992 Aug 1;89(15):6698–6702. [PMC free article] [PubMed]
  • Kerjaschki D, Ullrich R, Diem K, Pietromonaco S, Orlando RA, Farquhar MG. Identification of a pathogenic epitope involved in initiation of Heymann nephritis. Proc Natl Acad Sci U S A. 1992 Dec 1;89(23):11179–11183. [PMC free article] [PubMed]
  • Furukawa T, Ozawa M, Huang RP, Muramatsu T. A heparin binding protein whose expression increases during differentiation of embryonal carcinoma cells to parietal endoderm cells: cDNA cloning and sequence analysis. J Biochem. 1990 Aug;108(2):297–302. [PubMed]
  • Nakamoto M, Ozawa M, Jacinto SD, Furukawa T, Natori Y, Shirahama H, Yonezawa S, Nakayama T, Muramatsu T. Mouse heparin binding protein-44 (HBP-44) associates with brushin, a high-molecular-weight glycoprotein antigen common to the kidney and teratocarcinomas. J Biochem. 1993 Sep;114(3):344–349. [PubMed]
  • Kounnas MZ, Argraves WS, Strickland DK. The 39-kDa receptor-associated protein interacts with two members of the low density lipoprotein receptor family, alpha 2-macroglobulin receptor and glycoprotein 330. J Biol Chem. 1992 Oct 15;267(29):21162–21166. [PubMed]
  • Raychowdhury R, Niles JL, McCluskey RT, Smith JA. Autoimmune target in Heymann nephritis is a glycoprotein with homology to the LDL receptor. Science. 1989 Jun 9;244(4909):1163–1165. [PubMed]
  • Kerjaschki D, Farquhar MG. Immunocytochemical localization of the Heymann nephritis antigen (GP330) in glomerular epithelial cells of normal Lewis rats. J Exp Med. 1983 Feb 1;157(2):667–686. [PMC free article] [PubMed]
  • Kerjaschki D, Miettinen A, Farquhar MG. Initial events in the formation of immune deposits in passive Heymann nephritis. gp330-anti-gp330 immune complexes form in epithelial coated pits and rapidly become attached to the glomerular basement membrane. J Exp Med. 1987 Jul 1;166(1):109–128. [PMC free article] [PubMed]
  • Bu G, Williams S, Strickland DK, Schwartz AL. Low density lipoprotein receptor-related protein/alpha 2-macroglobulin receptor is an hepatic receptor for tissue-type plasminogen activator. Proc Natl Acad Sci U S A. 1992 Aug 15;89(16):7427–7431. [PMC free article] [PubMed]
  • Orth K, Madison EL, Gething MJ, Sambrook JF, Herz J. Complexes of tissue-type plasminogen activator and its serpin inhibitor plasminogen-activator inhibitor type 1 are internalized by means of the low density lipoprotein receptor-related protein/alpha 2-macroglobulin receptor. Proc Natl Acad Sci U S A. 1992 Aug 15;89(16):7422–7426. [PMC free article] [PubMed]
  • Willnow TE, Goldstein JL, Orth K, Brown MS, Herz J. Low density lipoprotein receptor-related protein and gp330 bind similar ligands, including plasminogen activator-inhibitor complexes and lactoferrin, an inhibitor of chylomicron remnant clearance. J Biol Chem. 1992 Dec 25;267(36):26172–26180. [PubMed]
  • Moestrup SK, Nielsen S, Andreasen P, Jørgensen KE, Nykjaer A, Røigaard H, Gliemann J, Christensen EI. Epithelial glycoprotein-330 mediates endocytosis of plasminogen activator-plasminogen activator inhibitor type-1 complexes. J Biol Chem. 1993 Aug 5;268(22):16564–16570. [PubMed]
  • Christensen EI, Gliemann J, Moestrup SK. Renal tubule gp330 is a calcium binding receptor for endocytic uptake of protein. J Histochem Cytochem. 1992 Oct;40(10):1481–1490. [PubMed]
  • Biemesderfer D, Dekan G, Aronson PS, Farquhar MG. Biosynthesis of the gp330/44-kDa Heymann nephritis antigenic complex: assembly takes place in the ER. Am J Physiol. 1993 Jun;264(6 Pt 2):F1011–F1020. [PubMed]
  • Herz J, Goldstein JL, Strickland DK, Ho YK, Brown MS. 39-kDa protein modulates binding of ligands to low density lipoprotein receptor-related protein/alpha 2-macroglobulin receptor. J Biol Chem. 1991 Nov 5;266(31):21232–21238. [PubMed]
  • Kounnas MZ, Chappell DA, Strickland DK, Argraves WS. Glycoprotein 330, a member of the low density lipoprotein receptor family, binds lipoprotein lipase in vitro. J Biol Chem. 1993 Jul 5;268(19):14176–14181. [PubMed]
  • Orlando RA, Farquhar MG. Identification of a cell line that expresses a cell surface and a soluble form of the gp330/receptor-associated protein (RAP) Heymann nephritis antigenic complex. Proc Natl Acad Sci U S A. 1993 May 1;90(9):4082–4086. [PMC free article] [PubMed]
  • Glenn KC, Carney DH, Fenton JW, 2nd, Cunningham DD. Thrombin active site regions required for fibroblast receptor binding and initiation of cell division. J Biol Chem. 1980 Jul 25;255(14):6609–6616. [PubMed]
  • Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. [PubMed]
  • Glabe CG, Harty PK, Rosen SD. Preparation and properties of fluorescent polysaccharides. Anal Biochem. 1983 Apr 15;130(2):287–294. [PubMed]
  • Mehlman T, Burgess WH. Detection and characterization of heparin-binding proteins with a gel overlay procedure. Anal Biochem. 1990 Jul;188(1):159–163. [PubMed]
  • Cardin AD, Weintraub HJ. Molecular modeling of protein-glycosaminoglycan interactions. Arteriosclerosis. 1989 Jan-Feb;9(1):21–32. [PubMed]
  • Busch SJ, Sassone-Corsi P. Dimers, leucine zippers and DNA-binding domains. Trends Genet. 1990 Feb;6(2):36–40. [PubMed]
  • Miettinen A, Törnroth T, Ekblom P, Virtanen I, Linder E. Nephritogenic and non-nephritogenic epithelial antigens in autoimmune and passive Heymann nephritis. Lab Invest. 1984 Apr;50(4):435–446. [PubMed]
  • Warshawsky I, Bu G, Schwartz AL. Identification of domains on the 39-kDa protein that inhibit the binding of ligands to the low density lipoprotein receptor-related protein. J Biol Chem. 1993 Oct 15;268(29):22046–22054. [PubMed]
  • Stow JL, Sawada H, Farquhar MG. Basement membrane heparan sulfate proteoglycans are concentrated in the laminae rarae and in podocytes of the rat renal glomerulus. Proc Natl Acad Sci U S A. 1985 May;82(10):3296–3300. [PMC free article] [PubMed]

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