Functional Changes in Scavenger Receptor Binding Conformation Are Induced by Charge Mutants Spanning the Entire Collagen Domain*
- From the Lipid Metabolism Unit and Nessel Gene Therapy Center, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts 02114
Abstract
Macrophage scavenger receptors are trimeric integral membrane proteins that bind a diverse array of negatively charged ligands. They have been shown to play a role in the pathogenesis of atherosclerosis and in host responses to microbial infections. Earlier mutational studies demonstrated that the distal segment of the collagen domain of the receptor was critically important for high affinity ligand binding activity. In this study, mutations spanning the entire collagen domain were generated and binding was assayed in transfected cells, as well as in assays employing a secreted, receptor fusion protein. Many of the distal, positively charged C-terminal residues in the type II collagen domain of the receptor, previously reported to be essential for binding at 37 °C, were found not to be critical for binding at 4 °C. Conversely, more proximally charged residues of the collagen receptor that have not been previously mutated were shown to have substantial effects on binding that were also temperature-dependent. These data suggest that scavenger receptor ligand recognition depends on more complex conformational interactions, involving charged residues throughout the entire collagen domain, than was previously recognized.
Footnotes
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↵* This work was supported in part by National Institutes of Health Grants DK50305 and HL 45098 and an American Heart Association Grant in Aid.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵‡ To whom correspondence should be addressed: Lipid Metabolism Unit and Nessel Gene Therapy Center, GRJ 1328, Massachusetts General Hospital and Harvard Medical School, Boston, MA 02114.
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↵1 The abbreviations used are: LDL, low density lipoprotein; SR, scavenger receptor(s); AcLDL, acetylated LDL; PBS, phosphate-buffered saline; BSA, bovine serum albumin; DAB, 3,3′-diaminobenzidine tetrahydrochloride.
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- Received October 27, 1997.
- Revision received May 11, 1998.
- The American Society for Biochemistry and Molecular Biology, Inc.
