Topology of scavenger receptor class B type I (SR-BI) on brush border membrane.
Both hydropathy plot and in vitro translation results predict the topology of SR-BI; the receptor is an integral membrane protein of 509 amino acids, consisting of a short cytoplasmic N-terminus of 9 amino acids followed by a first transmembrane domain of 22 amino acids, the extracellular domain of 408 amino acids, the second transmembrane domain of 22 amino acids, and the cytoplasmic C-terminus of 47 amino acids. The immunoblot of rBBMV in the presence or absence of pAb589 peptide antigen (the C-terminal 22 amino acid residues of SR-BI) confirmed that the bands at apparent molecular weight of 140 and 210 kDa are SR-BI related protein which might be multimeric forms of SR-BI. 125I apo A-I overlay analysis showed that SR-BI can bind to its ligand, apo A-I, only when it is thoroughly matured--glycosylated and dimerized. The antibody which was generated against extracellular domain of SR-BI (pAb230) not only prevented 125I-labeled apo A-I from binding to 140 kDa band but also inhibited the esterified cholesterol uptake of rabbit BBMV with its IC50 value of 40 microg/ml of IgG. In contrast, the antibody generated against the C-terminal domain of SR-BI (pAb589) did not show any effect either on cholesterol uptake of rabbit BBMV or 125I-labeled apo A-I binding to 140 kDa band. Overall results show that the ligand binding site of SR-BI in rabbit BBMV is located in extracellular domain, and SR-BI is only functional when it is part of dimeric forms which rationalize the previously found cooperative nature of the binding interaction and maybe a fundamental finding towards the so far poorly understood mechanism of SR-BI function.
Han
Chang-Hoon
CH
Brain Korea 21, School of Agricultural Biotechnology, Seoul National University, 441-744 Suwon, Korea.
Lee
Mun-Han
MH
D000595:Amino Acid Sequence; D000818:Animals; D016632:Apolipoprotein A-I / Q000378:metabolism; D001665:Binding Sites / Q000502:physiology; D015153:Blotting, Western; D018955:CD36 Antigens / Q000378:metabolism*; D018938:Caco-2 Cells; D002788:Cholesterol Esters / Q000378:metabolism; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism; D007421:Intestine, Small / Q000378:metabolism* / Q000648:ultrastructure; D007457:Iodine Radioisotopes; D008565:Membrane Proteins / Q000378:metabolism*; D008871:Microvilli / Q000378:metabolism; D008969:Molecular Sequence Data; D011817:Rabbits; D011971:Receptors, Immunologic; D018110:Receptors, Lipoprotein / Q000378:metabolism*; D051116:Receptors, Scavenger; D051122:Scavenger Receptors, Class B; D013499:Surface Properties
D016632:Apolipoprotein A-I; D018955:CD36 Antigens; D002788:Cholesterol Esters; D007457:Iodine Radioisotopes; D008565:Membrane Proteins; D011971:Receptors, Immunologic; D018110:Receptors, Lipoprotein; D051116:Receptors, Scavenger; C495124:SCARB1 protein, human; C454739:Scarb1 protein, mouse; D051122:Scavenger Receptors, Class B