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| Hauptseite > Publikationsdatenbank > Nanodiscs allow the use of integral membrane proteins as analytes in surface plasmon resonance studies |
| Hauptseite > Publikationsdatenbank > Nanodiscs allow the use of integral membrane proteins as analytes in surface plasmon resonance studies |
| Journal Article | PreJuSER-11819 |
; ;
2011
Elsevier
San Diego, Calif.
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Please use a persistent id in citations: doi:10.1016/j.ab.2010.08.028
Abstract: Nanodiscs are small-sized and flat model membranes that provide a close to native environment for reconstitution of integral membrane proteins. Incorporation of membrane proteins into nanodiscs results in water-soluble proteolipid particles making the membrane proteins amenable to a multitude of bioanalytical techniques originally developed for soluble proteins. The transmembrane domain of the human CD4 receptor was fused to ubiquitin with a preceding N-terminal decahistidine tag. The resulting integral membrane protein was incorporated into nanodiscs. Binding of the nanodisc-inserted histidine-tagged protein to a monoclonal anti-pentahistidine antibody was quantified using surface plasmon resonance (SPR) experiments. For the first time, a membrane-inserted transmembrane protein was employed as analyte while the antibody served as ligand immobilized on the sensor chip surface. SPR experiments were conducted in single-cycle mode. We demonstrate that the nanodisc-incorporated membrane protein showed nearly identical affinity toward the antibody as did the soluble decahistidine-tagged ubiquitin studied in a comparative experiment. Advantages of the new experimental setup and potential applications are discussed.
Keyword(s): Antibodies, Monoclonal: immunology (MeSH) ; Antigens, CD4: chemistry (MeSH) ; Antigens, CD4: genetics (MeSH) ; Antigens, CD4: metabolism (MeSH) ; Biosensing Techniques: methods (MeSH) ; Humans (MeSH) ; Kinetics (MeSH) ; Membrane Proteins: chemistry (MeSH) ; Membrane Proteins: genetics (MeSH) ; Membrane Proteins: metabolism (MeSH) ; Nanostructures: chemistry (MeSH) ; Oligopeptides: chemistry (MeSH) ; Oligopeptides: genetics (MeSH) ; Oligopeptides: metabolism (MeSH) ; Protein Binding (MeSH) ; Recombinant Fusion Proteins: chemistry (MeSH) ; Recombinant Fusion Proteins: genetics (MeSH) ; Recombinant Fusion Proteins: metabolism (MeSH) ; Surface Plasmon Resonance: methods (MeSH) ; Ubiquitin: chemistry (MeSH) ; Ubiquitin: genetics (MeSH) ; Ubiquitin: metabolism (MeSH) ; Antibodies, Monoclonal ; Antigens, CD4 ; Membrane Proteins ; Oligopeptides ; Recombinant Fusion Proteins ; Ubiquitin ; J ; Surface plasmon resonance (auto) ; Biacore (auto) ; Single-cycle kinetics (auto) ; Nanodisc (auto)
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