title: Capturing RNA-dependent pathways for cryo-EM analysis.
abstract: Cryo-Electron Microscopy (EM) is a powerful technique to visualize biological processes at nanometer resolution. Structural studies of macromolecular assemblies are typically performed on individual complexes that are biochemically isolated from their cellular context. Here we present a molecular imaging platform to capture and view multiple components of cellular pathways within a functionally relevant framework. We utilized the bacterial protein synthesis machinery as a model system to develop our approach. By using modified Affinity Grid surfaces, we were able to recruit multiple protein assemblies bound to nascent strands of mRNA. The combined use of Affinity Capture technology and single particle electron microscopy provide the basis for visualizing RNA-dependent pathways in a remarkable new way.
authors.affiliation: Virginia Tech Carilion Research Institute, Roanoke, VA, 24016, USA.
authors.forename: Justin R
authors.initials: JR
authors.lastname: Tanner
authors.affiliation: Department of Biomedical Engineering, University of Virginia, Charlottesville, VA, 22908, USA.
authors.forename: Katherine
authors.initials: K
authors.lastname: Degen
authors.affiliation: Virginia Tech Carilion Research Institute, Roanoke, VA, 24016, USA.
authors.forename: Brian L
authors.initials: BL
authors.lastname: Gilmore
authors.affiliation: Virginia Tech Carilion Research Institute, Roanoke, VA, 24016, USA.
authors.forename: Deborah F
authors.initials: DF
authors.lastname: Kelly
keywords: Affinity Capture technology; Lipid monolayer; Protein synthesis; Transcription