title: Affinity grid-based cryo-EM of PKC binding to RACK1 on the ribosome.
abstract: Affinity grids (AG) are specialized EM grids that bind macromolecular complexes containing tagged proteins to obtain maximum occupancy for structural analysis through single-particle EM. In this study, utilizing AG, we show that His-tagged activated PKC βII binds to the small ribosomal subunit (40S). We reconstructed a cryo-EM map which shows that PKC βII interacts with RACK1, a seven-bladed β-propeller protein present on the 40S and binds in two different regions close to blades 3 and 4 of RACK1. This study is a first step in understanding the molecular framework of PKC βII/RACK1 interaction and its role in translation.
authors.affiliation: Department of Biochemistry and Molecular Biophysics, Columbia University, New York City, NY 10032, USA.
authors.forename: Gyanesh
authors.initials: G
authors.lastname: Sharma
authors.forename: Jesper
authors.initials: J
authors.lastname: Pallesen
authors.forename: Sanchaita
authors.initials: S
authors.lastname: Das
authors.forename: Robert
authors.initials: R
authors.lastname: Grassucci
authors.forename: Robert
authors.initials: R
authors.lastname: Langlois
authors.forename: Cheri M
authors.initials: CM
authors.lastname: Hampton
authors.forename: Deborah F
authors.initials: DF
authors.lastname: Kelly
authors.forename: Amedee
authors.initials: A
authors.lastname: des Georges
authors.forename: Joachim
authors.initials: J
authors.lastname: Frank
mesh_terms: D020285:Cryoelectron Microscopy / Q000295:instrumentation / Q000379:methods*; D019204:GTP-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D008958:Models, Molecular; D009363:Neoplasm Proteins / Q000737:chemistry* / Q000378:metabolism; D014176:Protein Biosynthesis / Q000502:physiology*; D011487:Protein Conformation; D011493:Protein Kinase C / Q000737:chemistry* / Q000378:metabolism; D000075370:Receptors for Activated C Kinase; D011956:Receptors, Cell Surface / Q000737:chemistry* / Q000378:metabolism; D054682:Ribosome Subunits, Small, Eukaryotic / Q000378:metabolism*
chemical_list: D009363:Neoplasm Proteins; C419368:RACK1 protein, human; D000075370:Receptors for Activated C Kinase; D011956:Receptors, Cell Surface; D011493:Protein Kinase C; D019204:GTP-Binding Proteins