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Issue 9, 2015
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Temperature-cycle microscopy reveals single-molecule conformational heterogeneity

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Our previous temperature-cycle study reported FRET transitions between different states on FRET-labeled polyprolines [Yuan et al., PCCP, 2011, 13, 1762]. The conformational origin of such transitions, however, was left open. In this work, we apply temperature-cycle microscopy of single FRET-labeled polyproline and dsDNA molecules and compare their responses to resolve the conformational origin of different FRET states. We observe different steady-state FRET distributions and different temperature-cycle responses in the two samples. Our temperature-cycle results on single molecules resemble the results in steady-state measurements but reveal a dark state which could not be observed otherwise. By comparing the timescales and probabilities of different FRET states in temperature-cycle traces, we assign the conformational heterogeneity reflected by different FRET states to linker dynamics, dye–chain and dye–dye interactions. The dark state and low-FRET state are likely due to dye–dye interactions at short separations.

Graphical abstract: Temperature-cycle microscopy reveals single-molecule conformational heterogeneity

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The article was received on 25 Nov 2014, accepted on 27 Jan 2015 and first published on 30 Jan 2015

Article type: Paper
DOI: 10.1039/C4CP05486E
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Citation: Phys. Chem. Chem. Phys., 2015,17, 6532-6544
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    Temperature-cycle microscopy reveals single-molecule conformational heterogeneity

    H. Yuan, A. Gaiduk, J. R. Siekierzycka, S. Fujiyoshi, M. Matsushita, D. Nettels, B. Schuler, C. A. M. Seidel and M. Orrit, Phys. Chem. Chem. Phys., 2015, 17, 6532
    DOI: 10.1039/C4CP05486E

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