Abstract:SUMMARY
The interaction of high density lipoprotein (HDL) with its receptor, scavenger receptor BI (SR-BI), is critical for lowering plasma cholesterol levels and reducing the risk for cardiovascular disease. The HDL/SR-BI complex facilitates delivery of cholesterol into cells and is likely mediated by receptor dimerization. This work describes the use of nuclear magnetic resonance (NMR) spectroscopy to generate the first high-resolution structure of the C-terminal transmembrane domain of SR-BI. This region of… Show more
“…Figure 1.Sequence conservation alignments and structural modeling of proline residues in SR-BI's C-terminal transmembrane domain and proximal extracellular domain.( A ) Multiple-species sequence alignments of SR-BI(405–475) were generated using the MUSCLE algorithm. ( B ) NMR structure of SR-BI(405–475) [18] (PDB: 5KTF) is shown, and the locations of conserved proline residues are highlighted as spheres. ( C ) A homology model of SR-BI's extracellular domain was generated using MODELLER [30] with conserved proline residues indicated.…”
Section: Resultsmentioning
confidence: 99%
“…Oligomerization of SR-BI receptors may play an important role in cholesterol transport [ 21 ], and dimerization is mediated, at least in part, by the N- and C-terminal transmembrane domains [ 18 , 23 , 24 ]. To determine if the presence of proline residues in the C-terminal transmembrane domain and adjacent extracellular domain is required for SR-BI's ability to oligomerize, we harvested cell lysates in buffer containing PFO (a non-dissociative detergent) [ 34 ], separated samples by PFO-PAGE, and performed immunoblot analysis using the antibody directed against the C-terminal region of SR-BI.…”
Section: Resultsmentioning
confidence: 99%
“…The use of structure–function studies has expanded our knowledge of SR-BI cholesterol transport functions [ 14 – 18 ]. Homology models of SR-BI's extracellular domain provided physical evidence of a hydrophobic cavity that can spatially accommodate cholesterol molecules [ 19 ].…”
Section: Introductionmentioning
confidence: 99%
“…We observed homo-dimerization of SR-BI's C-terminal region in live cells using fluorescence resonance energy transfer experiments [ 24 ]. Dimerization of the C-terminal region is likely mediated by multiple interactions and may include a potential glycine dimerization motif as well as a leucine zipper dimerization motif that partially spans the C-terminal transmembrane domain [ 18 ]. Mutation of all leucine residues within the leucine zipper motif to alanine decreased receptor oligomerization and SR-BI-mediated cholesterol transport [ 18 ].…”
Section: Introductionmentioning
confidence: 99%
“…Seeking to gain additional insights into how the C-terminal region of SR-BI contributes to its function, we solved the high-resolution NMR structure of an SR-BI peptide encompassing residues 405–475 [SR-BI(405–475)], which spans the C-terminal transmembrane domain and proximal residues in the extracellular domain [ 18 ]. The amino acid sequence and NMR structure of SR-BI(405–475) revealed four conserved proline (Pro/P) residues, including one proline (Pro-459) that induces a kink in the transmembrane α-helix.…”
High-density lipoproteins (HDLs) facilitate reverse cholesterol transport, a process in which HDL removes cholesterol from circulation and carries it to the liver for biliary excretion. Reverse cholesterol transport is also facilitated by HDL's high-affinity receptor, scavenger receptor-BI (SR-BI), by mechanisms that are not fully understood. To improve our understanding of SR-BI function, we previously solved the NMR (nuclear magnetic resonance) structure of a peptide encompassing amino acids 405–475 of SR-BI. This segment of SR-BI, that includes the functionally critical C-terminal transmembrane domain and part of the extracellular domain, also contains four conserved proline (Pro) residues. We hypothesized that these proline residues support SR-BI in a conformation that allows for efficient cholesterol transport. To test this, we generated individual Pro-to-alanine mutations in full-length SR-BI and transiently expressed the mutant receptors in COS-7 cells to measure the effects on SR-BI-mediated cholesterol transport functions. Our findings reveal that HDL cell association and uptake of HDL-cholesteryl esters are impaired by mutation of Pro-412, Pro-438, or the transmembrane proline kink residue (Pro-459). In addition, SR-BI-mediated cholesterol efflux and membrane cholesterol distribution are impaired by mutation of Pro-412 or Pro-438, indicating that these residues are essential for a fully functional SR-BI receptor. Furthermore, we demonstrate that Pro-408 is necessary for proper SR-BI expression, but mutation of Pro-408 does not cause SR-BI to become misfolded or rapidly degraded by the proteasome or the lysosome. We conclude that key proline residues play an important role in SR-BI function by allowing for the efficient transport of cholesterol between cells and HDL.
“…Figure 1.Sequence conservation alignments and structural modeling of proline residues in SR-BI's C-terminal transmembrane domain and proximal extracellular domain.( A ) Multiple-species sequence alignments of SR-BI(405–475) were generated using the MUSCLE algorithm. ( B ) NMR structure of SR-BI(405–475) [18] (PDB: 5KTF) is shown, and the locations of conserved proline residues are highlighted as spheres. ( C ) A homology model of SR-BI's extracellular domain was generated using MODELLER [30] with conserved proline residues indicated.…”
Section: Resultsmentioning
confidence: 99%
“…Oligomerization of SR-BI receptors may play an important role in cholesterol transport [ 21 ], and dimerization is mediated, at least in part, by the N- and C-terminal transmembrane domains [ 18 , 23 , 24 ]. To determine if the presence of proline residues in the C-terminal transmembrane domain and adjacent extracellular domain is required for SR-BI's ability to oligomerize, we harvested cell lysates in buffer containing PFO (a non-dissociative detergent) [ 34 ], separated samples by PFO-PAGE, and performed immunoblot analysis using the antibody directed against the C-terminal region of SR-BI.…”
Section: Resultsmentioning
confidence: 99%
“…The use of structure–function studies has expanded our knowledge of SR-BI cholesterol transport functions [ 14 – 18 ]. Homology models of SR-BI's extracellular domain provided physical evidence of a hydrophobic cavity that can spatially accommodate cholesterol molecules [ 19 ].…”
Section: Introductionmentioning
confidence: 99%
“…We observed homo-dimerization of SR-BI's C-terminal region in live cells using fluorescence resonance energy transfer experiments [ 24 ]. Dimerization of the C-terminal region is likely mediated by multiple interactions and may include a potential glycine dimerization motif as well as a leucine zipper dimerization motif that partially spans the C-terminal transmembrane domain [ 18 ]. Mutation of all leucine residues within the leucine zipper motif to alanine decreased receptor oligomerization and SR-BI-mediated cholesterol transport [ 18 ].…”
Section: Introductionmentioning
confidence: 99%
“…Seeking to gain additional insights into how the C-terminal region of SR-BI contributes to its function, we solved the high-resolution NMR structure of an SR-BI peptide encompassing residues 405–475 [SR-BI(405–475)], which spans the C-terminal transmembrane domain and proximal residues in the extracellular domain [ 18 ]. The amino acid sequence and NMR structure of SR-BI(405–475) revealed four conserved proline (Pro/P) residues, including one proline (Pro-459) that induces a kink in the transmembrane α-helix.…”
High-density lipoproteins (HDLs) facilitate reverse cholesterol transport, a process in which HDL removes cholesterol from circulation and carries it to the liver for biliary excretion. Reverse cholesterol transport is also facilitated by HDL's high-affinity receptor, scavenger receptor-BI (SR-BI), by mechanisms that are not fully understood. To improve our understanding of SR-BI function, we previously solved the NMR (nuclear magnetic resonance) structure of a peptide encompassing amino acids 405–475 of SR-BI. This segment of SR-BI, that includes the functionally critical C-terminal transmembrane domain and part of the extracellular domain, also contains four conserved proline (Pro) residues. We hypothesized that these proline residues support SR-BI in a conformation that allows for efficient cholesterol transport. To test this, we generated individual Pro-to-alanine mutations in full-length SR-BI and transiently expressed the mutant receptors in COS-7 cells to measure the effects on SR-BI-mediated cholesterol transport functions. Our findings reveal that HDL cell association and uptake of HDL-cholesteryl esters are impaired by mutation of Pro-412, Pro-438, or the transmembrane proline kink residue (Pro-459). In addition, SR-BI-mediated cholesterol efflux and membrane cholesterol distribution are impaired by mutation of Pro-412 or Pro-438, indicating that these residues are essential for a fully functional SR-BI receptor. Furthermore, we demonstrate that Pro-408 is necessary for proper SR-BI expression, but mutation of Pro-408 does not cause SR-BI to become misfolded or rapidly degraded by the proteasome or the lysosome. We conclude that key proline residues play an important role in SR-BI function by allowing for the efficient transport of cholesterol between cells and HDL.
Purpose of Review
The binding of high-density lipoprotein (HDL) to its primary receptor, scavenger receptor class B type 1 (SR-B1), is critical for lowering plasma cholesterol levels and reducing cardiovascular disease risk. This review provides novel insights into how the structural elements of SR-B1 drive efficient function with an emphasis on bidirectional cholesterol transport.
Recent Findings
We have generated a new homology model of full-length human SR-B1 based on the recent resolution of the partial structures of other class B scavenger receptors. Interrogating this model against previously published observations allows us to generate structurally informed hypotheses about SR-B1’s ability to mediate HDL-cholesterol (HDL-C) transport. Furthermore, we provide a structural perspective as to why human variants of SR-B1 may result in impaired HDL-C clearance.
Summary
A comprehensive understanding of SR-B1’s structure–function relationships is critical to the development of therapeutic agents targeting SR-B1 and modulating cardiovascular disease risk.
Purpose of Review
As both a cholesterol acceptor and carrier in the reverse cholesterol transport (RCT) pathway, high-density lipoprotein (HDL) is putatively atheroprotective. However, current pharmacological therapies to increase plasma HDL cholesterol (HDL-c) concentration have paradoxically failed to prevent or reduce atherosclerosis and cardiovascular disease (CVD). Given that free cholesterol (FC) transfer between surfaces of lipoproteins and cells is reversible, excess plasma FC can be transferred to the cells of peripheral tissue sites resulting in atherosclerosis. Here, we summarize potential mechanisms contributing to this paradox and highlight the role of excess free cholesterol (FC) bioavailability in atherosclerosis vs. atheroprotection.
Recent Findings
Recent findings have established a complex relationship between HDL-c concentration and atherosclerosis. Systemic scavenger receptor class B type 1 (SR-B1) knock out (KO) mice exhibit with increased diet-induced atherosclerosis despite having an elevated plasma HDL-c concentration compared to wild type (WT) mice. The greater bioavailability of HDL-FC in SR-B1 vs. WT mice is associated with a higher FC content in multiple cell types and tissue sites. These results suggest that dysfunctional HDL with high FC bioavailability is atheroprone despite high HDL-c concentration.
Summary
Past oversimplification of HDL-c involvement in cholesterol transport has led to the failures in HDL targeted therapy. Evidence suggests that FC-mediated functionality of HDL is of higher importance than its quantity; as a result, deciphering the regulatory mechanisms by which HDL-FC bioavailability can induce atherosclerosis can have far-reaching clinical implications.
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