title: Multifrequency Force Microscopy of Helical Protein Assembly on a Virus.
abstract: High-resolution microscopy techniques have been extensively used to investigate the structure of soft, biological matter at the nanoscale, from very thin membranes to small objects, like viruses. Electron microscopy techniques allow for obtaining extraordinary resolution by averaging signals from multiple identical structures. In contrast, atomic force microscopy (AFM) collects data from single entities. Here, it is possible to finely modulate the interaction with the samples, in order to be sensitive to their top surface, avoiding mechanical deformations. However, most biological surfaces are highly curved, such as fibers or tubes, and ultimate details of their surface are in the vicinity of steep height variations. This limits lateral resolution, even when sharp probes are used. We overcome this problem by using multifrequency force microscopy on a textbook example, the Tobacco Mosaic Virus (TMV). We achieved unprecedented resolution in local maps of amplitude and phase shift of the second excited mode, recorded together with sample topography. Our data, which combine multifrequency imaging and Fourier analysis, confirm the structure deduced from averaging techniques (XRD, cryoEM) for surface features of single virus particles, down to the helical pitch of the coat protein subunits, 2.3 nm. Remarkably, multifrequency AFM images do not require any image postprocessing.
authors.affiliation: CIC nanoGUNE, Tolosa Hiribidea 76, E-20018 San Sebastián-Donostia, Spain.
authors.forename: Annalisa
authors.initials: A
authors.lastname: Calò
authors.affiliation: CIC nanoGUNE, Tolosa Hiribidea 76, E-20018 San Sebastián-Donostia, Spain.
authors.forename: Aitziber
authors.initials: A
authors.lastname: Eleta-Lopez
authors.affiliation: CIC nanoGUNE, Tolosa Hiribidea 76, E-20018 San Sebastián-Donostia, Spain.
authors.forename: Pablo
authors.initials: P
authors.lastname: Stoliar
authors.affiliation: CIC nanoGUNE, Tolosa Hiribidea 76, E-20018 San Sebastián-Donostia, Spain.|Ikerbasque, Basque Foundation for Science, Maria Diaz de Haro 3, 48013 Bilbao, Spain.
authors.forename: David
authors.initials: D
authors.lastname: De Sancho
authors.affiliation: Institute Center for Energy (iEnergy), Masdar Institute of Science and Technology, Abu Dhabi, United Arab Emirates.
authors.forename: Sergio
authors.initials: S
authors.lastname: Santos
authors.affiliation: Catalan Institute of Nanoscience and Nanotechnology (ICN2), CSIC and the Barcelona Institute of Science and Technology, Campus UAB, Bellaterra, 08193 Barcelona, Spain.
authors.forename: Albert
authors.initials: A
authors.lastname: Verdaguer
authors.affiliation: CIC nanoGUNE, Tolosa Hiribidea 76, E-20018 San Sebastián-Donostia, Spain.|Ikerbasque, Basque Foundation for Science, Maria Diaz de Haro 3, 48013 Bilbao, Spain.
authors.forename: Alexander M
authors.initials: AM
authors.lastname: Bittner
mesh_terms: D018625:Microscopy, Atomic Force / Q000379:methods*; D014027:Tobacco Mosaic Virus / Q000648:ultrastructure*; D014771:Virion / Q000648:ultrastructure*