title: Phosphoinositide-incorporated lipid-protein nanodiscs: A tool for studying protein-lipid interactions.
abstract: Phosphatidylinositol (PtdIns) is phosphorylated at D-3, D-4, and/or D-5 of the inositol ring to produce seven distinct lipid second messengers known as phosphoinositides (PIs). The PI level is temporally and spatially controlled at the cytosolic face of the cellular membrane. Effectors containing PI-binding domains (e.g., PH, PX, FYVE, ENTH, FERM) associate with specific PIs. This process is crucial for the localization of a variety of cell-signaling proteins, thereby regulating intracellular membrane trafficking, cell growth and survival, cytoskeletal organization, and so on. However, quantitative assessments of protein-PI interactions are generally difficult due to insolubility of PIs in aqueous solution. Here we incorporated PIs into a lipid-protein nanoscale bilayer (nanodisc), which is applied for studying the protein-PI interactions using pull-down binding assay, fluorescence polarization, and nuclear magnetic resonance studies, each facilitating fast, quantitative, and residue-specific evaluation of the protein-PI interactions. Therefore, the PI-incorporated nanodisc could be used as a versatile tool for studying the protein-lipid interactions by various biochemical and biophysical techniques.
authors.affiliation: Department of Structural Biology, Faculty of Advanced Life Science, Hokkaido University, N-21, W-11, Kita-ku, Sapporo 001-0021, Japan.
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Kobashigawa
authors.forename: Kohsuke
authors.initials: K
authors.lastname: Harada
authors.forename: Naoki
authors.initials: N
authors.lastname: Yoshida
authors.forename: Kenji
authors.initials: K
authors.lastname: Ogura
authors.forename: Fuyuhiko
authors.initials: F
authors.lastname: Inagaki
mesh_terms: D016632:Apolipoprotein A-I / Q000737:chemistry / Q000378:metabolism; D002462:Cell Membrane / Q000737:chemistry* / Q000378:metabolism*; D005454:Fluorescence Polarization; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008051:Lipid Bilayers / Q000737:chemistry / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008297:Male; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D008958:Models, Molecular; D036103:Nanotechnology / Q000379:methods*; D010716:Phosphatidylinositols / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity; D014867:Water / Q000737:chemistry
chemical_list: D016632:Apolipoprotein A-I; D008051:Lipid Bilayers; D008565:Membrane Proteins; D010716:Phosphatidylinositols; D014867:Water