Molecular dissection of ligand binding sites on the low density lipoprotein receptor-related protein

Item Type:	Article
Title:	Molecular dissection of ligand binding sites on the low density lipoprotein receptor-related protein
Creators Name:	Willnow, T.E. and Orth, K. and Herz, J.
Abstract:	The low density lipoprotein receptor-related protein (LRP) is a large multifunctional receptor that is involved in the cellular uptake of a number of functionally diverse ligands including apoE-rich remnant lipoproteins, lipoprotein lipase, alpha 2-macroglobulin-protease complexes, plasminogen activator-inhibitor complexes, and the active protease tissue-type plasminogen activator. Ligand binding and competition experiments suggest that most LRP ligands bind to specific, independent sites on the large 515-kDa subunit of the receptor. In a previous study (Moestrup, S.K., Holtet, T.L., Etzerodt, M., Thogersen, H.C., Nykjaer, A., Andreasen, P.A., Rasmussen, H.H., Sottrup-Jensen, L., and Gliemann, J. (1993) J. Biol. Chem. 268, 13691-13696), ligand blotting was used to localize the binding sites for urokinase-type plasminogen activator-plasminogen activator inhibitor-1 (PAI-1) complexes and for alpha 1-macroglobulin to a proteolytic fragment of LRP containing the second cluster of complement-type cysteine-rich repeats. Here, we have used a recombinant DNA approach to express functionally restricted chimeric "LRP-minireceptors" containing two different regions of the extracellular domain of the receptor in cultured cells. Receptor-associated protein, a negative modulator of LRP activity, is bound and internalized by cells transfected with either construct. A minireceptor containing the cluster of eight complement-type cysteine-rich repeats followed by four epidermal growth factor precursor homologous domains binds and internalizes 125I-labeled plasminogen activator-PAI-1 complexes. It also mediates the cellular uptake of the uncomplexed protease tissue-type plasminogen activator (tPA), suggesting that the tPA and PAI-1 binding sites on LRP are in close vicinity and might promote cooperative binding of tPA-PAI-1 complexes. However, alpha 2-macroglobulin is not internalized by this minireceptor suggesting that this ligand requires the presence of a single epidermal growth factor-repeat which is contained in the previously studied proteolytic fragment but is absent from the minireceptor.
Keywords:	Base Sequence, Binding Sites, CHO Cells, Cell Membrane, DNA Primers, Kinetics, LDL-Receptor Related Protein 1, Macromolecular Substances, Structural Models, Molecular Sequence Data, Plasminogen Activator Inhibitor 1, Polymerase Chain Reaction, Protein Sorting Signals, Secondary Protein Structure, Immunologic Receptors, Transfection, Urinary Plasminogen Activator, alpha-Macroglobulins, Animals, Cricetinae
Source:	Journal of Biological Chemistry
ISSN:	0021-9258
Publisher:	American Society for Biochemistry and Molecular Biology
Volume:	269
Number:	22
Page Range:	15827-15832
Date:	3 June 1994
Official Publication:	https://doi.org/10.1016/S0021-9258(17)40755-1
PubMed:	View item in PubMed

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