title: Direct binding of cholesterol to the purified membrane region of SCAP: mechanism for a sterol-sensing domain.
abstract: Mammalian cells control their membrane composition by regulating the vesicular transport of membrane-bound sterol regulatory element binding proteins (SREBPs) from endoplasmic reticulum (ER) to Golgi. Transport is blocked by cholesterol, which triggers SCAP, the SREBP escort protein, to bind to Insigs, which are ER retention proteins. The cholesterol trigger mechanism is unknown. Using recombinant SCAP purified in detergent, we show that cholesterol acts by binding with high affinity and specificity to the 767 amino acid octahelical membrane region of SCAP. This octahelical region contains a conserved pentahelical sterol-sensing domain found in six other polytopic membrane proteins. We show that the membrane domain of SCAP is a tetramer and that cholesterol binding is inhibited by cationic amphiphiles, raising the possibility of allosteric regulation by positively charged phospholipids. The current studies show that cells control their cholesterol content through receptor-ligand interactions and not through changes in the physical properties of the membrane.
authors.affiliation: Department of Molecular Genetics, University of Texas Southwestern Medical Center, Dallas 75390, USA.
authors.forename: Arun
authors.initials: A
authors.lastname: Radhakrishnan
authors.forename: Li-Ping
authors.initials: LP
authors.lastname: Sun
authors.forename: Hyock Joo
authors.initials: HJ
authors.lastname: Kwon
authors.forename: Michael S
authors.initials: MS
authors.lastname: Brown
authors.forename: Joseph L
authors.initials: JL
authors.lastname: Goldstein
mesh_terms: D000494:Allosteric Regulation; D000818:Animals; D001665:Binding Sites; D002462:Cell Membrane / Q000378:metabolism*; D002784:Cholesterol / Q000378:metabolism*; D006224:Cricetinae; D003902:Detergents / Q000378:metabolism; D004331:Drosophila melanogaster / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins / Q000378:metabolism*; D010743:Phospholipids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012243:Rhodopsin / Q000378:metabolism; D012680:Sensitivity and Specificity; D013261:Sterols / Q000378:metabolism
chemical_list: D003902:Detergents; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D010743:Phospholipids; D011994:Recombinant Proteins; C102196:SREBP cleavage-activating protein; D013261:Sterols; D012243:Rhodopsin; D002784:Cholesterol