title: Glass is a Viable Substrate for Precision Force Microscopy of Membrane Proteins.
abstract: Though ubiquitous in optical microscopy, glass has long been overlooked as a specimen supporting surface for high resolution atomic force microscopy (AFM) investigations due to its roughness. Using bacteriorhodopsin from Halobacterium salinarum and the translocon SecYEG from Escherichia coli, we demonstrate that faithful images of 2D crystalline and non-crystalline membrane proteins in lipid bilayers can be obtained on microscope cover glass following a straight-forward cleaning procedure. Direct comparison between AFM data obtained on glass and on mica substrates show no major differences in image fidelity. Repeated association of the ATPase SecA with the cytoplasmic protrusion of SecYEG demonstrates that the translocon remains competent for binding after tens of minutes of continuous AFM imaging. This opens the door for precision long-timescale investigations of the active translocase in near-native conditions and, more generally, for integration of high resolution biological AFM with many powerful optical techniques that require non-birefringent substrates.
authors.affiliation: Department of Physics and Astronomy, University of Missouri-Columbia, Columbia, Missouri 65211 USA.
authors.forename: Nagaraju
authors.initials: N
authors.lastname: Chada
authors.affiliation: Department of Physics and Astronomy, University of Missouri-Columbia, Columbia, Missouri 65211 USA.
authors.forename: Krishna P
authors.initials: KP
authors.lastname: Sigdel
authors.affiliation: Department of Physics and Astronomy, University of Missouri-Columbia, Columbia, Missouri 65211 USA.
authors.forename: Raghavendar Reddy Sanganna
authors.initials: RR
authors.lastname: Gari
authors.affiliation: Department of Physics and Astronomy, University of Missouri-Columbia, Columbia, Missouri 65211 USA.
authors.forename: Tina Rezaie
authors.initials: TR
authors.lastname: Matin
authors.affiliation: Department of Biochemistry, University of Missouri-Columbia, Columbia, Missouri 65211 USA.
authors.forename: Linda L
authors.initials: LL
authors.lastname: Randall
authors.affiliation: 1] Department of Physics and Astronomy, University of Missouri-Columbia, Columbia, Missouri 65211 USA [2] Department of Biochemistry, University of Missouri-Columbia, Columbia, Missouri 65211 USA.
authors.forename: Gavin M
authors.initials: GM
authors.lastname: King
mesh_terms: D000251:Adenosine Triphosphatases / Q000032:analysis / Q000737:chemistry; D000538:Aluminum Silicates; D001426:Bacterial Proteins / Q000032:analysis / Q000737:chemistry; D001436:Bacteriorhodopsins / Q000032:analysis / Q000737:chemistry; D029968:Escherichia coli Proteins / Q000032:analysis / Q000737:chemistry; D005898:Glass; D019617:Halobacterium salinarum / Q000737:chemistry; D007091:Image Processing, Computer-Assisted; D008051:Lipid Bilayers / Q000737:chemistry; D008565:Membrane Proteins / Q000032:analysis* / Q000737:chemistry; D026901:Membrane Transport Proteins / Q000032:analysis / Q000737:chemistry; D018625:Microscopy, Atomic Force / Q000295:instrumentation / Q000379:methods*; D000069816:SEC Translocation Channels; D000081416:SecA Proteins
chemical_list: D000538:Aluminum Silicates; D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D008051:Lipid Bilayers; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D000069816:SEC Translocation Channels; D001436:Bacteriorhodopsins; D000251:Adenosine Triphosphatases; D000081416:SecA Proteins; C011934:mica