Number of the records: 1
Environmentally sensitive probes for monitoring protein-membrane interactions at nanomolar concentrations
- 1.
SYSNO ASEP 0475308 Document Type J - Journal Article R&D Document Type Journal Article Subsidiary J Článek ve WOS Title Environmentally sensitive probes for monitoring protein-membrane interactions at nanomolar concentrations Author(s) Shvadchak, Volodymyr V. (UOCHB-X) ORCID, RID
Kucherak, Oleksandr (UOCHB-X)
Afitska, Kseniia (UOCHB-X) ORCID, RID
Dziuba, D. (DE)
Yushchenko, Dmytro A. (UOCHB-X) ORCID, RIDSource Title Biochimica Et Biophysica Acta-Biomembranes. - : Elsevier - ISSN 0005-2736
Roč. 1859, č. 5 (2017), s. 852-859Number of pages 8 s. Language eng - English Country NL - Netherlands Keywords solvatochromic probes ; fluorescence ; protein-membrane interaction ; affinity ; binding stoichiometry ; alpha-synuclein Subject RIV CE - Biochemistry OECD category Biochemistry and molecular biology Institutional support UOCHB-X - RVO:61388963 UT WOS 000398870800020 EID SCOPUS 85012115798 DOI 10.1016/j.bbamem.2017.01.021 Annotation Solvatochromic probes are suitable tools for quantitative characterization of protein-membrane interactions. Based on diverse fluorophores these probes have different fluorescent properties and therefore demonstrate different responses when applied for sensing the interactions of biomolecules. Surprisingly, to the best of our knowledge, no systematic comparison of the sensitivities of solvatochromic dyes for monitoring protein-membrane interactions was described. Hence, a rational choice of an optimal environmentally sensitive probe for such experiments is usually not a straightforward task. In this work we developed a series of thiol-reactive fluorescent probes based on the fluorophores with high sensitivity to their environment and compared them with two widely used DNS and DMN probes. We investigated the responses of these probes to the interaction of probe-labeled presynaptic protein alpha-synuclein with lipid membranes. We observed that newly synthesized probes based on fluorene and chromone dyes, which combine the strongest brightness and significant changes of fluorescence intensity, demonstrated the highest sensitivity to interaction of alpha-synuclein with lipid membranes. They are especially beneficial for sensing in scattering media such as solutions of lipid vesicles. We show that the described probes permit quantitative measurements of ot-synuclein binding to lipid membranes at low nanomolar concentrations. We developed a detailed protocol for measuring Kd and binding stoichiometry for interaction of soluble peripheral proteins with membranes based on the response of the environmentally sensitive fluorescent probes. We applied this protocol for quantification of the affinity of alpha-synuclein to anionic membranes and found that it is substantially higher than it was earlier reported. Workplace Institute of Organic Chemistry and Biochemistry Contact asep@uochb.cas.cz ; Kateřina Šperková, Tel.: 232 002 584 ; Viktorie Chládková, Tel.: 232 002 434 Year of Publishing 2018
Number of the records: 1