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DC Field | Value | Language |
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dc.contributor.author | Caorsi, V | - |
dc.contributor.author | Ushakov, DS | - |
dc.contributor.author | West, TG | - |
dc.contributor.author | Setta-Kaffetzi, N | - |
dc.contributor.author | Ferenczi, MA | - |
dc.date.accessioned | 2022-01-29T17:29:53Z | - |
dc.date.available | 2022-01-29T17:29:53Z | - |
dc.date.issued | 2010-09-02 | - |
dc.identifier.citation | Caorsi, V., Ushakov, D.S., West, T.G., Setta-Kaffetzi, N. and Ferenczi, M.A. (2011) 'FRET characterisation for cross-bridge dynamics in single-skinned rigor muscle fibres', European Biophysics Journal, 40 (1), pp. 13 - 27. doi: 10.1007/s00249-010-0624-9. | en_US |
dc.identifier.issn | 0175-7571 | - |
dc.identifier.uri | https://bura.brunel.ac.uk/handle/2438/24028 | - |
dc.description.abstract | Copyright © The Author(s) 2010. In this work we demonstrate for the first time the use of Förster resonance energy transfer (FRET) as an assay to monitor the dynamics of cross-bridge conformational changes directly in single muscle fibres. The advantage of FRET imaging is its ability to measure distances in the nanometre range, relevant for structural changes in actomyosin cross-bridges. To reach this goal we have used several FRET couples to investigate different locations in the actomyosin complex. We exchanged the native essential light chain of myosin with a recombinant essential light chain labelled with various thiol-reactive chromophores. The second fluorophore of the FRET couple was introduced by three approaches: labelling actin, labelling SH1 cysteine and binding an adenosine triphosphate (ATP) analogue. We characterise FRET in rigor cross-bridges: in this condition muscle fibres are well described by a single FRET population model which allows us to evaluate the true FRET efficiency for a single couple and the consequent donor–acceptor distance. The results obtained are in good agreement with the distances expected from crystallographic data. The FRET characterisation presented herein is essential before moving onto dynamic measurements, as the FRET efficiency differences to be detected in an active muscle fibre are on the order of 10–15% of the FRET efficiencies evaluated here. This means that, to obtain reliable results to monitor the dynamics of cross-bridge conformational changes, we had to fully characterise the system in a steady-state condition, demonstrating firstly the possibility to detect FRET and secondly the viability of the present approach to distinguish small FRET variations. | en_US |
dc.description.sponsorship | Royal Society (Newton International fellowship to V. Caorsi) and Milstein Fund, Medical Research Council (G0601747). | en_US |
dc.format.extent | 13 - 27 | - |
dc.format.medium | Print-Electronic | - |
dc.language | en | - |
dc.language.iso | en_US | en_US |
dc.publisher | Springer Science and Business Media LLC | en_US |
dc.rights | Copyright © The Author(s) 2010.Open Access. This is an open access article distributed under the terms of the Creative Commons Attribution Noncommercial License (https://creativecommons.org/licenses/by-nc/2.0), which permits any noncommercial use, distribution, and reproduction in any medium, provided the original author(s) and source are credited. | - |
dc.rights.uri | https://creativecommons.org/licenses/by-nc/2.0 | - |
dc.subject | FRET | en_US |
dc.subject | acceptor photobleaching | en_US |
dc.subject | spectral analysis | en_US |
dc.subject | FLIM | en_US |
dc.subject | rigor muscle fibre | en_US |
dc.subject | actomyosin interactions | en_US |
dc.title | FRET characterisation for cross-bridge dynamics in single-skinned rigor muscle fibres | en_US |
dc.type | Article | en_US |
dc.identifier.doi | https://doi.org/10.1007/s00249-010-0624-9 | - |
dc.relation.isPartOf | European Biophysics Journal | - |
pubs.issue | 1 | - |
pubs.publication-status | Published | - |
pubs.volume | 40 | - |
dc.identifier.eissn | 1432-1017 | - |
Appears in Collections: | Dept of Health Sciences Research Papers |
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