Affinity grid-based cryo-EM of PKC binding to RACK1 on the ribosome

J Struct Biol. 2013 Feb;181(2):190-4. doi: 10.1016/j.jsb.2012.11.006. Epub 2012 Dec 8.

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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cryoelectron Microscopy / instrumentation
  • Cryoelectron Microscopy / methods*
  • GTP-Binding Proteins / chemistry*
  • GTP-Binding Proteins / metabolism
  • Humans
  • Models, Molecular*
  • Neoplasm Proteins / chemistry*
  • Neoplasm Proteins / metabolism
  • Protein Biosynthesis / physiology*
  • Protein Conformation*
  • Protein Kinase C / chemistry*
  • Protein Kinase C / metabolism
  • Receptors for Activated C Kinase
  • Receptors, Cell Surface / chemistry*
  • Receptors, Cell Surface / metabolism
  • Ribosome Subunits, Small, Eukaryotic / metabolism*

Substances

  • Neoplasm Proteins
  • RACK1 protein, human
  • Receptors for Activated C Kinase
  • Receptors, Cell Surface
  • Protein Kinase C
  • GTP-Binding Proteins