Highly purified scavenger receptor class B, type I reconstituted into phosphatidylcholine/cholesterol liposomes mediates high affinity high density lipoprotein binding and selective lipid uptake.
The murine class B, type I scavenger receptor mSR-BI is a high and low density lipoprotein (HDL and LDL) receptor that mediates selective uptake of cholesteryl esters. Here we describe a reconstituted phospholipid/cholesterol liposome assay of the binding and selective uptake activities of SR-BI derived from detergent-solubilized cells. The assay, employing lysates from epitope-tagged receptor (mSR-BI-t1)-expressing mammalian and insect cells, recapitulated many features of SR-BI activity in intact cells, including high affinity and saturable (125)I-HDL binding, selective lipid uptake from [(3)H]cholesteryl ether-labeled HDL, and poor inhibition of HDL receptor activity by LDL. The novel properties of a mutated receptor (Q402R/Q418R, normal LDL binding but loss of most HDL binding) were reproduced in the assay, as was the ability of the SR-BI homologue CD36 to bind HDL but not mediate efficient lipid uptake. In this assay, essentially homogeneously pure mSR-BI-t1, prepared by single-step immunoaffinity chromatography, mediated high affinity HDL binding and efficient selective lipid uptake from HDL. Thus, SR-BI-mediated HDL binding and selective lipid uptake are intrinsic properties of the receptor that do not require the intervention of other proteins or specific cellular structures or compartments.
Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
Bin
B
Liu
Monty
M
Krieger
D000818:Animals; D018955:CD36 Antigens / Q000302:isolation & purification / Q000378:metabolism / Q000502:physiology*; D019556:COS Cells; D002784:Cholesterol / Q000378:metabolism; D002846:Chromatography, Affinity; D006801:Humans; D050356:Lipid Metabolism; D008075:Lipoproteins, HDL / Q000378:metabolism*; D008077:Lipoproteins, LDL / Q000378:metabolism; D008081:Liposomes / Q000378:metabolism*; D008565:Membrane Proteins; D010713:Phosphatidylcholines / Q000378:metabolism; D011971:Receptors, Immunologic; D018110:Receptors, Lipoprotein; D051116:Receptors, Scavenger; D051122:Scavenger Receptors, Class B; D018411:Spodoptera; D014162:Transfection
D018955:CD36 Antigens; D008075:Lipoproteins, HDL; D008077:Lipoproteins, LDL; D008081:Liposomes; D008565:Membrane Proteins; D010713:Phosphatidylcholines; D011971:Receptors, Immunologic; D018110:Receptors, Lipoprotein; D051116:Receptors, Scavenger; C495124:SCARB1 protein, human; C454739:Scarb1 protein, mouse; D051122:Scavenger Receptors, Class B; D002784:Cholesterol