Glycine Dimerization Motif in the N-terminal Transmembrane Domain of the High Density Lipoprotein Receptor SR-BI Required for Normal Receptor Oligomerization and Lipid Transport

Abstract

Scavenger receptor class B, type I (SR-BI), a CD36 superfamily member, is an oligomeric high density lipoprotein (HDL) receptor that mediates negatively cooperative HDL binding and selective lipid uptake. We identified in the N-terminal transmembrane (N-TM) domain of SR-BI a conserved glycine dimerization motif, G 15 X 2 G 18 X 3 AX 2 G 25 , of which the submotif G 18 X 3 AX 2 G 25 significantly contributes to homodimerization and lipid uptake activity. SR-BI variants were generated by mutations (single or multiple Gly 3 Leu substitutions) or by replacing the N-TM domain with those from other CD36 superfamily members containing (croquemort) or lacking (lysosomal integral membrane protein (LIMP) II) this glycine motif (chimeras). None of the SR-BI variants exhibited altered surface expression (based on antibody binding) or HDL binding. However, the G15L/G18L/G25L triple mutant exhibited reductions in cell surface homo-oligomerization (>10-fold) and the rate of selective lipid uptake (ϳ2-fold). Gly 18 and Gly 25 were necessary for normal lipid uptake activity of SR-BI and the SR-BI/ croquemort chimera. The lipid uptake activity of the glycine motif-deficient SR-BI/LIMP II chimera was low but could be increased by introducing glycines at positions 18 and 25. The rate of lipid uptake mediated by SR-BI/LIMP II chimeras was proportional to the extent of receptor oligomerization. Thus, the glycine dimerization motif G 18 X 3 AX 2 G 25 in the N-TM domain of SR-BI contributes substantially to the homo-oligomerization and lipid transport activity of SR-BI but does not influence the negative cooperativity of HDL binding. Oligomerization-independent binding cooperativity suggests that classic allostery is not involved and that the negative cooperativity is probably the consequence of a "lattice effect" (interligand steric interference accompanying binding to adjacent receptors).The high density lipoprotein (HDL) receptor SR-BI 2 is a cell surface receptor expressed in the liver and other tissues that regulates plasma lipoprotein cholesterol levels and metabolism (Refs. 1 and 3-6; for a review, see Ref.2). SR-BI mediates cellular uptake of cholesteryl esters and other lipids from HDL and has been proposed to play a key role in moving cholesterol from peripheral tissues to the liver and then out of the body (Refs. 7-10; for a review, see Ref.2). SR-BI influences HDL metabolism in humans (11,12) in whom the risk of atherosclerotic disease is inversely or directly proportional to plasma concentrations of HDL cholesterol and low density lipoprotein (LDL) cholesterol, respectively (13-15). Studies in mice have shown that SR-BI protects against atherosclerosis and coronary heart disease (Refs. 16 -22; for a review, see Ref.2).SR-BI is a member of the CD36 superfamily of proteins in metazoans (ϳ25-30% sequence identities). Members of this superfamily include lysosomal integral membrane protein (LIMP) II (23) and croquemort (a Drosophila melanogaster hemocyte/macrophage receptor for apoptotic cells) (24). Most CD36 superfamily membe...

Exoplasmic cysteine Cys384 of the HDL receptor SR-BI is critical for its sensitivity to a small-molecule inhibitor and normal lipid transport activity

“…The phenotype of the C384S mutant is reminiscent of that of SR-BI with a mutagenic disruption of the glycine dimerization motif (GX 3 AX 2 G) in its N-terminal transmembrane domain: essentially normal surface expression and HDL binding, but a twofold reduction in lipid uptake (24). There is almost a complete loss of receptor homooligomerization when the dimerization motif is disrupted, as detected by cross-linking with the watersoluble, membrane-impermeable cross-linker bis(sulfosuccinimidyl)suberate (BS 3 ) or by coimmunoprecipitation.…”

“…Mutagenesis and genetics studies have identified a few residues in SR-BI that play roles in lipoprotein binding and lipid transport (24)(25)(26)(27)(28) and in interactions with HCV (29). For example, disruption of the glycine dimerization motif interferes with oligomerization and efficient lipid transfer into cells (24). However, the molecular mechanisms underlying SR-BI's activities are not well defined (7).…”

“…SR-BI's homooligomerization (20)(21)(22)(23)(24) is mediated by a glycine dimerization motif in its N-terminal transmembrane domain. Mutagenesis and genetics studies have identified a few residues in SR-BI that play roles in lipoprotein binding and lipid transport (24)(25)(26)(27)(28) and in interactions with HCV (29). For example, disruption of the glycine dimerization motif interferes with oligomerization and efficient lipid transfer into cells (24).…”

Critical residues and motifs for homodimerization of the first transmembrane domain of the plasma membrane glycoprotein CD36

“…However, the underlying molecular mechanisms of signal transduction of CD36 have remained elusive. A recent report showed that the first transmembrane domain (TM1) of scavenger receptor class B, type I (SR-BI), a homolog of CD36, is critically involved in its oligomerization and function (24). Small-residue motif 15 GXXGXXXAXXG 25 in TM1 of SR-BI was identified to be critical to receptor homo-oligomerization and ligand binding via mutagenesis analysis.…”

“…Gaidukov et al (24) recently identified a conserved smallresidue motif 18 GXXXAXXG 25 , which significantly contributes to the homodimerization of SR-BI. This motif corresponds to 16 GXXXAXXG 23 sequence in CD36 TM1.…”