Action-FRET: Probing the Molecular Conformation of Mass-Selected Gas-Phase Peptides with Forster Resonance Energy Transfer Detected by Acceptor-Specific Fragmentation

Abstract : The use of Forster resonance energy transfer (FRET) as a probe of the structure of biological molecules through fluorescence measurements in solution is well-attested. The transposition of this technique to the gas phase is appealing since it opens the perspective of combining the structural accuracy of FRET with the specificity and selectivity of mass spectrometry (MS). Here, we report FRET results on gas-phase polyalanine ions obtained by measuring FRET efficiency through specific photofragmentation rather than fluorescence. The structural sensitivity of the method was tested using commercially available chromophores (QSY 7 and carboxyrhodamine 575) grafted on a series of small, alanine-based peptides of differing sizes. The photofragmentation of these systems was investigated through action spectroscopy, and their conformations were probed using ion mobility spectrometry (IMS) and Monte Carlo minimization (MCM) simulations. We show that specific excitation of the donor chromophore results in the observation of fragments that are specific to the electronic excitation of the acceptor chromophore. This shows that energy transfer took place between the two chromophores and hence that the action-FRET technique can be used as a new and sensitive probe of the structure of gas-phase biomolecules, which opens perspectives as a new tool in structural biology.
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Submitted on : Thursday, October 2, 2014 - 11:06:40 AM
Last modification on : Thursday, March 15, 2018 - 10:30:28 AM

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Steven Daly, Frédéric Poussigue, Anne-Laure Simon, Luke Macaleese, Franck Bertorelle, et al.. Action-FRET: Probing the Molecular Conformation of Mass-Selected Gas-Phase Peptides with Forster Resonance Energy Transfer Detected by Acceptor-Specific Fragmentation. Analytical Chemistry, American Chemical Society, 2014, 86 (17), pp.8798-8804. ⟨10.1021/ac502027y⟩. ⟨hal-01070732⟩

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