Fig 1: ORP1-ORD transfers cholesterol but not PIPs. a Schematic diagram of human ORP1L and ORP1S. The structural elements in ORD are colored as follows: N-terminal loop (blue), lid (magenta), core (yellow). The color scheme of ORP1-ORD is consistent throughout the schematic representations unless indicated. b Lipid-binding analyses for ORP1-ORD. c DHE transport assays without or with PIP in the acceptor liposomes. Shown at the bottom are the initial velocities of DHE transport subtracted by that without ORP1. d PIP transport assays with DHE-containing acceptor liposomes. (For all bar graphs, data are shown as mean ± s.e.m. (error bar), n = 3)
Fig 2: PI(3,4)P2/PI(4,5)P2 stimulates cholesterol transport by ORP1L in vitro. a SDS-PAGE of full-length ORP1L. b Lipid-binding analyses for ORP1L and its PH Domain. c, d DHE transport assays for ORP1-ORD (c) and ORP1L (d) with PIP embedded in the donor liposomes. Shown at the bottom are the initial velocities. e PI(4,5)P2 (left) and PI4P (right) transport assays for ORP1L. (For all bar graphs, data are shown as mean ± s.e.m. (error bar), n = 3)
Fig 3: PI(3,4)P2/PI(4,5)P2 allosterically stimulates ORP1 function. a Lysosomal cholesterol release assay in ORP1L KO cells overexpressing ORP1L mutants (scale bar = 10 μm). b Quantitation of LELs filipin intensities of transfected cells in a (data are shown as mean ± s.d. (error bar); n = 5–15). c Working model for PI(4,5)P2/PI(3,4)P2-stimulated cholesterol transport by ORP1-ORD. The red arrows indicate the orientations of lipid-binding tunnel to membrane
Fig 4: Mutational analysis of the cholesterol binding site on ORP1-ORD. a DHE-binding assays of ORP1-ORD mutants. b PI(3,4)P2-stimulated DHE transport assays of ORP1-ORD mutations with both lipids embedded in donor liposomes. (Data are shown as mean ± s.e.m. (error bar), n = 3)
Fig 5: Structures of cholesterol-bound ORP1-ORD in open conformations. a, b Overall structures of ORP1-ORD in complex with cholesterol at 2.6 Å (a) and at 3.4 Å (b). The cholesterol ligand is highlighted as green stick. c Detailed binding mode of cholesterol in the 2.6 Å structure. For clarity, part of the structure has been removed to show the ligand-binding tunnel. Dashed line, polar interaction
Supplier Page from Abcam for Anti-ORP1 antibody [EPR8646]