title: Detection of metal binding sites on functional S-layer nanoarrays using single molecule force spectroscopy.
abstract: Crystalline bacterial cell surface layers (S-layers) show the ability to recrystallize into highly regular pattern on solid supports. In this study, the genetically modified S-layer protein SbpA of Lysinibacillus sphaericus CCM 2177, carrying a hexa-histidine tag (His(6)-tag) at the C-terminus, was used to generate functionalized two-dimensional nanoarrays on a silicon surface. Atomic force microscopy (AFM) was applied to explore the topography and the functionality of the fused His(6)-tags. The accessibility of the His(6)-tags was demonstrated by in-situ anti-His-tag antibody binding to the functional S-layer array. The metal binding properties of the His(6)-tag was investigated by single molecule force microscopy. For this purpose, newly developed tris-NTA was tethered to the AFM tips via a flexible polyethylene glycol (PEG) linker. The functionalized tips showed specific interactions with S-layer containing His(6)-tags in the presence of nickel ions. Thus the His(6)-tag is located at the outer surface of the S-layer and can be used for stable but reversible attachment of functional tris-NTA derivatives.
authors.affiliation: Institute of Biophysics, Johannes Kepler University of Linz, Austria.
authors.forename: Jilin
authors.initials: J
authors.lastname: Tang
authors.forename: Andreas
authors.initials: A
authors.lastname: Ebner
authors.forename: Bernhard
authors.initials: B
authors.lastname: Kraxberger
authors.forename: Michael
authors.initials: M
authors.lastname: Leitner
authors.forename: Alba
authors.initials: A
authors.lastname: Hykollari
authors.forename: Christian
authors.initials: C
authors.lastname: Kepplinger
authors.forename: Christian
authors.initials: C
authors.lastname: Grunwald
authors.forename: Hermann J
authors.initials: HJ
authors.lastname: Gruber
authors.forename: Robert
authors.initials: R
authors.lastname: Tampé
authors.forename: Uwe B
authors.initials: UB
authors.lastname: Sleytr
authors.forename: Nicola
authors.initials: N
authors.lastname: Ilk
authors.forename: Peter
authors.initials: P
authors.lastname: Hinterdorfer
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry; D001665:Binding Sites; D008562:Membrane Glycoproteins / Q000737:chemistry*; D018625:Microscopy, Atomic Force; D036103:Nanotechnology / Q000379:methods*
chemical_list: D001426:Bacterial Proteins; D008562:Membrane Glycoproteins; C055764:S-layer proteins