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Issue 13, 2011
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Unfolding dynamics of cytochrome c revealed by single-molecule and ensemble-averaged spectroscopy

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Abstract

Denaturant-induced conformational change of yeast iso-1-cytochrome c (Cytc) has been comprehensively investigated in the single-molecule and bulk phases. By fluorescence-quenching experiments with dye-labelled heme-protein (Alexa 488-labelled Cytc, Cytc-A488), we clearly show that the fluorescence quenching observed from folded Cytc-A488 is due mainly to photoinduced electron transfer (PET) between electron-donating amino acids such as tryptophan and the dye attached to the protein. In addition, the unfolding process of Cytc-A488 observed in the single-molecule and bulk phases can be explained well in terms of a three-state model: Cytc unfolds through an intermediate with a native-like compactness. By quantitative analysis of fluorescence correlation spectroscopy (FCS) data, we were able to observe a relaxation time of ∼1.5 μs corresponding to segmental motion and fast folding dynamics of 55 μs in the unfolded state of Cytc. The results presented here also suggest that a combination of single-molecule and ensemble-averaged spectroscopy is necessary to provide convincing and comprehensive assignments of protein kinetics.

Graphical abstract: Unfolding dynamics of cytochrome c revealed by single-molecule and ensemble-averaged spectroscopy

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Publication details

The article was received on 29 Nov 2010, accepted on 18 Jan 2011 and first published on 09 Feb 2011


Article type: Paper
DOI: 10.1039/C0CP02689A
Citation: Phys. Chem. Chem. Phys., 2011,13, 5651-5658
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    Unfolding dynamics of cytochrome c revealed by single-molecule and ensemble-averaged spectroscopy

    J. Choi, S. Kim, T. Tachikawa, M. Fujitsuka and T. Majima, Phys. Chem. Chem. Phys., 2011, 13, 5651
    DOI: 10.1039/C0CP02689A

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