Protein-protein interactions in reversibly assembled nanopatterns.
We describe herein a platform to study protein-protein interactions and to form functional protein complexes in nanoscopic surface domains. For this purpose, we employed multivalent chelator (MCh) templates, which were fabricated in a stepwise procedure combining dip-pen nanolithography (DPN) and molecular recognition-directed assembly. First, we demonstrated that an atomic force microscope (AFM) tip inked with an oligo(ethylene glycol) (OEG) disulfide compound bearing terminal biotin groups can be used to generate biotin patterns on gold achieving line widths below 100 nm, a generic platform for fabrication of functional nanostructures via the highly specific biotin-streptavidin recognition. Subsequently, we converted such biotin/streptavidin patterns into functional MCh patterns for reversible assembly of histidine-tagged (His-tagged) proteins via the attachment of a tris-nitriloacetic acid (trisNTA) biotin derivative. Fluorescence microscopy confirmed reversible immobilization of the receptor subunit ifnar2-His10 and its interaction with interferon-alpha2 labeled with fluorescent quantum dots in a 7 x 7 dot array consisting of trisNTA spots with a diameter of approximately 230 nm. Moreover, we carried out characterization of the specificity, stability, and reversibility as well as quantitative real-time analysis of protein-protein interactions at the fabricated nanopatterns by imaging surface plasmon resonance. Our work offers a route for construction and analysis of functional protein-based nanoarchitectures.
Rakickas
Tomas
T
Department of Functional Nanomaterials, Institute of Physics, Savanoriu 231, LT-02300 Vilnius, Lithuania.
Gavutis
Martynas
M
Reichel
Annett
A
Piehler
Jacob
J
Liedberg
Bo
B
Valiokas
Ramūnas
R
D001710:Biotin / Q000737:chemistry; D002614:Chelating Agents / Q000494:pharmacology; D019855:Ethylene Glycol / Q000737:chemistry; D007700:Kinetics; D018625:Microscopy, Atomic Force; D008856:Microscopy, Fluorescence; D008956:Models, Chemical; D053758:Nanoparticles / Q000737:chemistry*; D049329:Nanostructures / Q000737:chemistry; D036103:Nanotechnology / Q000379:methods*; D025941:Protein Interaction Mapping; D011506:Proteins / Q000737:chemistry*; D020349:Surface Plasmon Resonance
D002614:Chelating Agents; D011506:Proteins; D001710:Biotin; D019855:Ethylene Glycol