A Peptide-Based, Ratiometric Biosensor Construct for Direct Fluorescence Detection of a Protein Analyte
| Autor: | Yves Mely, Alexander P. Demchenko, Karin Enander, A. Linnéa Olsson, Daniel Kanmert, Danièle Altschuh, Andrey S. Klymchenko, Dmytro A. Yushchenko, Laurence Choulier |
|---|---|
| Přispěvatelé: | Laboratoire de Biophotonique et Pharmacologie - UMR 7213 (LBP), Réseau nanophotonique et optique, Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Université de Haute-Alsace (UHA) Mulhouse - Colmar (Université de Haute-Alsace (UHA))-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Université de Haute-Alsace (UHA) Mulhouse - Colmar (Université de Haute-Alsace (UHA))-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Réseau nanophotonique et optique, Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Université de Haute-Alsace (UHA) Mulhouse - Colmar (Université de Haute-Alsace (UHA))-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Université de Haute-Alsace (UHA) Mulhouse - Colmar (Université de Haute-Alsace (UHA)) |
| Jazyk: | angličtina |
| Rok vydání: | 2008 |
| Předmět: |
Analyte
Fluorophore [SDV]Life Sciences [q-bio] Molecular Sequence Data Biomedical Engineering Pharmaceutical Science Bioengineering Peptide Biosensing Techniques 02 engineering and technology 010402 general chemistry 01 natural sciences Immunoglobulin Fab Fragments chemistry.chemical_compound Amino Acid Sequence ComputingMilieux_MISCELLANEOUS Fluorescent Dyes Pharmacology chemistry.chemical_classification Binding Sites Chemistry C-terminus Organic Chemistry Proteins 021001 nanoscience & nanotechnology Tautomer Fluorescence 0104 chemical sciences 3. Good health Dissociation constant Kinetics Spectrometry Fluorescence Biochemistry Biophysics Peptides 0210 nano-technology Biosensor Biotechnology |
| Zdroj: | Bioconjugate Chemistry Bioconjugate Chemistry, American Chemical Society, 2008, 19 (9), pp.1864-1870. ⟨10.1021/bc800159d⟩ |
| ISSN: | 1043-1802 1520-4812 |
| Popis: | We present the design, synthesis, and functional evaluation of peptide-based fluorescent constructs for wavelength-ratiometric biosensing of a protein analyte. The concept was shown using the high-affinity model interaction between the 18 amino acid peptide pTMVP and a recombinant antibody fragment, Fab57P. pTMVP was functionalized in two different positions with 6-bromomethyl-2-(2-furanyl)-3-hydroxychromone, an environmentally sensitive fluorophore with a two-band emission. The equilibrium dissociation constant of the interaction between pTMVP and Fab57P was largely preserved upon labeling. The biosensor ability of the labeled peptide constructs was evaluated in terms of the relative intensity change of the emission bands from the normal (N*) and tautomer (T*) excited-state species of the fluorophore ( I(N*)/I(T*)) upon binding of Fab57P. When the peptide was labeled in the C terminus, the I(N*)/I(T*) ratio changed by 40% upon analyte binding, while labeling close to the residues most important for binding resulted in a construct that completely lacked ratiometric biosensor ability. Integrated biosensor elements for reagentless detection, where peptides and ratiometric fluorophores are combined to ensure robustness in both recognition and signaling, are expected to become an important contribution to the design of future protein quantification assays in immobilized formats. |
| Databáze: | OpenAIRE |
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