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. 1986 Feb 1;233(3):683–690. doi: 10.1042/bj2330683

Immunoprecipitation of the low-density-lipoprotein (LDL) receptor and its precursor from human monocyte-derived macrophages.

A K Soutar, B L Knight
PMCID: PMC1153086  PMID: 3707518

Abstract

Synthesis of the low-density-lipoprotein (LDL) receptor protein by cultured human monocyte-derived macrophages was demonstrated by immunoprecipitation of [35S]methionine-labelled cell extracts with a monoclonal antibody to the bovine adrenal LDL receptor. Although the antibody does not bind to or inhibit binding of 125I-LDL to the LDL receptor on intact fibroblasts, it specifically binds to a protein in extracts of human skin fibroblasts, of Mr approx. 130,000 under non-reducing conditions, that is able to bind LDL. In monocyte-derived macrophages, as in fibroblasts, the receptor is synthesized as a low-Mr precursor that is converted into the mature protein. The half-life of the precursor in human macrophages is approx. 44 min. In cells from two homozygous familial-hypercholesterolaemic subjects, only the precursor form of the receptor is synthesized. Detection of abnormalities of LDL-receptor synthesis in human mononuclear cells may be a useful aid in diagnosis of familial hypercholesterolaemia that is simpler and quicker than methods requiring growth of cultured skin fibroblasts.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Beisiegel U., Schneider W. J., Brown M. S., Goldstein J. L. Immunoblot analysis of low density lipoprotein receptors in fibroblasts from subjects with familial hypercholesterolemia. J Biol Chem. 1982 Nov 10;257(21):13150–13156. [PubMed] [Google Scholar]
  2. Beisiegel U., Schneider W. J., Goldstein J. L., Anderson R. G., Brown M. S. Monoclonal antibodies to the low density lipoprotein receptor as probes for study of receptor-mediated endocytosis and the genetics of familial hypercholesterolemia. J Biol Chem. 1981 Nov 25;256(22):11923–11931. [PubMed] [Google Scholar]
  3. Bruck C., Portetelle D., Glineur C., Bollen A. One-step purification of mouse monoclonal antibodies from ascitic fluid by DEAE Affi-Gel blue chromatography. J Immunol Methods. 1982 Sep 30;53(3):313–319. doi: 10.1016/0022-1759(82)90178-8. [DOI] [PubMed] [Google Scholar]
  4. Gavigan S. J., Knight B. L. The effects of low-density lipoprotein and cholesterol on acyl-coenzyme A: cholesterol acyltransferase activity in membranes from cultured human fibroblasts. Biochem J. 1983 Oct 15;216(1):93–100. doi: 10.1042/bj2160093. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Goldstein J. L., Basu S. K., Brunschede G. Y., Brown M. S. Release of low density lipoprotein from its cell surface receptor by sulfated glycosaminoglycans. Cell. 1976 Jan;7(1):85–95. doi: 10.1016/0092-8674(76)90258-0. [DOI] [PubMed] [Google Scholar]
  6. Goldstein J. L., Brown M. S. The LDL receptor locus and the genetics of familial hypercholesterolemia. Annu Rev Genet. 1979;13:259–289. doi: 10.1146/annurev.ge.13.120179.001355. [DOI] [PubMed] [Google Scholar]
  7. Knight B. L., Soutar A. K. Degradation by cultured fibroblasts and macrophages of unmodified and 1,2-cyclohexanedione-modified low-density lipoprotein from normal and homozygous familial hypercholesterolaemic subjects. Biochem J. 1982 Jan 15;202(1):145–152. doi: 10.1042/bj2020145. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  9. Schneider W. J., Beisiegel U., Goldstein J. L., Brown M. S. Purification of the low density lipoprotein receptor, an acidic glycoprotein of 164,000 molecular weight. J Biol Chem. 1982 Mar 10;257(5):2664–2673. [PubMed] [Google Scholar]
  10. Shechter I., Fogelman A. M., Haberland M. E., Seager J., Hokom M., Edwards P. A. The metabolism of native and malondialdehyde-altered low density lipoproteins by human monocyte-macrophages. J Lipid Res. 1981 Jan;22(1):63–71. [PubMed] [Google Scholar]
  11. Soutar A. K., Knight B. L. Degradation by cultured monocyte-derived macrophages from normal and familial hypercholesterolaemic subjects of modified and unmodified low-density lipoproteins. Biochem J. 1982 May 15;204(2):549–556. doi: 10.1042/bj2040549. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Terhorst C., van Agthoven A., LeClair K., Snow P., Reinherz E., Schlossman S. Biochemical studies of the human thymocyte cell-surface antigens T6, T9 and T10. Cell. 1981 Mar;23(3):771–780. doi: 10.1016/0092-8674(81)90441-4. [DOI] [PubMed] [Google Scholar]
  13. Tolleshaug H., Goldstein J. L., Schneider W. J., Brown M. S. Posttranslational processing of the LDL receptor and its genetic disruption in familial hypercholesterolemia. Cell. 1982 Oct;30(3):715–724. doi: 10.1016/0092-8674(82)90276-8. [DOI] [PubMed] [Google Scholar]
  14. Tolleshaug H., Hobgood K. K., Brown M. S., Goldstein J. L. The LDL receptor locus in familial hypercholesterolemia: multiple mutations disrupt transport and processing of a membrane receptor. Cell. 1983 Mar;32(3):941–951. doi: 10.1016/0092-8674(83)90079-x. [DOI] [PubMed] [Google Scholar]
  15. Traber M. G., Kayden H. J. Low density lipoprotein receptor activity in human monocyte-derived macrophages and its relation to atheromatous lesions. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5466–5470. doi: 10.1073/pnas.77.9.5466. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Wade D. P., Knight B. L., Soutar A. K. Detection of the low-density-lipoprotein receptor with biotin-low-density lipoprotein. A rapid new method for ligand blotting. Biochem J. 1985 Aug 1;229(3):785–790. doi: 10.1042/bj2290785. [DOI] [PMC free article] [PubMed] [Google Scholar]

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