title: A novel method to couple electrophysiological measurements and fluorescence imaging of suspended lipid membranes: the example of T5 bacteriophage DNA ejection.
abstract: We present an innovative method to couple electrophysiological measurements with fluorescence imaging of functionalized suspended bilayers. Our method combines several advantages: it is well suited to study transmembrane proteins that are difficult to incorporate in suspended bilayers, it allows single molecule resolution both in terms of electrophysiological measurements and fluorescence imaging, and it enables mechanical stimulations of the membrane. The approach comprises of two steps: first the reconstitution of membrane proteins in giant unilamellar vesicles; then the formation of a suspended bilayer spanning a 5 to 15 micron-wide aperture that can be visualized by high NA microscope objectives. We exemplified how the technique can be used to detect in real time the translocation of T5 DNA across the bilayer during its ejection from the bacteriophage capsid.
authors.affiliation: ESPCI Paristech, CNRS, Paris, France ; Aurélien Roux Lab, Biochemistry Department, University of Geneva, Geneva, Switzerland.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Chiaruttini
authors.affiliation: Institut de Biochimie et Biophysique Moléculaire et Cellulaire, Université Paris Sud-11, CNRS, Orsay, France.
authors.forename: Lucienne
authors.initials: L
authors.lastname: Letellier
authors.affiliation: ESPCI Paristech, CNRS, Paris, France ; Aurélien Roux Lab, Biochemistry Department, University of Geneva, Geneva, Switzerland ; Institut de Biochimie et Biophysique Moléculaire et Cellulaire, Université Paris Sud-11, CNRS, Orsay, France ; MechanoBiology Institute of Singapore, Singapore, Singapore.
authors.forename: Virgile
authors.initials: V
authors.lastname: Viasnoff
mesh_terms: D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure*; D004279:DNA, Viral / Q000378:metabolism; D055724:Electrophysiological Phenomena / Q000502:physiology*; D029968:Escherichia coli Proteins / Q000378:metabolism; D008051:Lipid Bilayers / Q000378:metabolism*; D008846:Micromanipulation; D061848:Optical Imaging / Q000379:methods*; D013604:T-Phages / Q000235:genetics; D057074:Virus Release / Q000502:physiology
chemical_list: D001425:Bacterial Outer Membrane Proteins; D004279:DNA, Viral; D029968:Escherichia coli Proteins; C050900:FhuA protein, E coli; D008051:Lipid Bilayers