title: Tuning membrane protein mobility by confinement into nanodomains.
abstract: High-speed atomic force microscopy (HS-AFM) can be used to visualize function-related conformational changes of single soluble proteins. Similar studies of single membrane proteins are, however, hampered by a lack of suitable flat, non-interacting membrane supports and by high protein mobility. Here we show that streptavidin crystals grown on mica-supported lipid bilayers can be used as porous supports for membranes containing biotinylated lipids. Using SecYEG (protein translocation channel) and GlpF (aquaglyceroporin), we demonstrate that the platform can be used to tune the lateral mobility of transmembrane proteins to any value within the dynamic range accessible to HS-AFM imaging through glutaraldehyde-cross-linking of the streptavidin. This allows HS-AFM to study the conformation or docking of spatially confined proteins, which we illustrate by imaging GlpF at sub-molecular resolution and by observing the motor protein SecA binding to SecYEG.
authors.affiliation: Center for Advanced Bioanalysis GmbH, Gruberstrasse 40-42, 4020 Linz, Austria.
authors.forename: Andreas
authors.initials: A
authors.lastname: Karner
authors.affiliation: Center for Advanced Bioanalysis GmbH, Gruberstrasse 40-42, 4020 Linz, Austria.
authors.forename: Benedikt
authors.initials: B
authors.lastname: Nimmervoll
authors.affiliation: Upper Austria University of Applied Sciences, Campus Linz, Garnisonstrasse 21, 4020 Linz, Austria.
authors.forename: Birgit
authors.initials: B
authors.lastname: Plochberger
authors.affiliation: EMBL Australia Node in Single Molecule Science and ARC Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, Sydney, Australia.
authors.forename: Enrico
authors.identifier: 0000-0002-7577-9042
authors.initials: E
authors.lastname: Klotzsch
authors.affiliation: Institute of Biophysics, Johannes Kepler University Linz, Gruberstrasse 40, 4020 Linz, Austria.
authors.forename: Andreas
authors.initials: A
authors.lastname: Horner
authors.affiliation: Institute of Biophysics, Johannes Kepler University Linz, Gruberstrasse 40, 4020 Linz, Austria.
authors.forename: Denis G
authors.initials: DG
authors.lastname: Knyazev
authors.affiliation: Institute of Biophysics, Johannes Kepler University Linz, Gruberstrasse 40, 4020 Linz, Austria.
authors.forename: Roland
authors.initials: R
authors.lastname: Kuttner
authors.affiliation: Institute of Biophysics, Johannes Kepler University Linz, Gruberstrasse 40, 4020 Linz, Austria.
authors.forename: Klemens
authors.initials: K
authors.lastname: Winkler
authors.affiliation: Institute of Biophysics, Johannes Kepler University Linz, Gruberstrasse 40, 4020 Linz, Austria.
authors.forename: Lukas
authors.initials: L
authors.lastname: Winter
authors.affiliation: Institute of Biophysics, Johannes Kepler University Linz, Gruberstrasse 40, 4020 Linz, Austria.
authors.forename: Christine
authors.initials: C
authors.lastname: Siligan
authors.affiliation: Institute of Biophysics, Johannes Kepler University Linz, Gruberstrasse 40, 4020 Linz, Austria.
authors.forename: Nicole
authors.initials: N
authors.lastname: Ollinger
authors.affiliation: Institute of Biophysics, Johannes Kepler University Linz, Gruberstrasse 40, 4020 Linz, Austria.
authors.forename: Peter
authors.identifier: 0000-0002-1792-2314
authors.initials: P
authors.lastname: Pohl
authors.affiliation: Center for Advanced Bioanalysis GmbH, Gruberstrasse 40-42, 4020 Linz, Austria.
authors.forename: Johannes
authors.initials: J
authors.lastname: Preiner
mesh_terms: D004926:Escherichia coli / Q000737:chemistry* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics; D008051:Lipid Bilayers / Q000737:chemistry*; D008565:Membrane Proteins / Q000737:chemistry*; D018625:Microscopy, Atomic Force / Q000379:methods; D000072417:Protein Domains; D021381:Protein Transport
chemical_list: D029968:Escherichia coli Proteins; D008051:Lipid Bilayers; D008565:Membrane Proteins