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Partial Donor-Donor Energy Migration (PDDEM) as a Fluorescence Spectroscopic Tool for Measuring Distances in Biomacromolecules
Umeå University, Faculty of Science and Technology, Chemistry.
Umeå University, Faculty of Science and Technology, Chemistry.
2002 (English)In: Spectrochimica Acta Part A - Molecular and Biomolecular Spectroscopy, ISSN 1386-1425, Vol. 58, no 5, 1087-1097 p.Article in journal (Refereed) Published
Abstract [en]

A theoretical model is presented, tested and applied for determining the rates of energy migration and distances within pairs of chemically identical fluorophores, so-called donors (D), which are exposed to different physical properties. The model is a general extension of the recently developed donor–donor energy migration (DDEM) model [J. Chem. Soc., Faraday Trans. 92 (1996)1563; J. Chem. Phys. 105 (1996) 10896] that applies to examining structure-function of biomacromolecules, such as proteins. Most fluorescent groups of the same kind incorporated at different positions (α and β) in a macromolecule exhibit shifts of the absorption and/or emission spectra, as well as different relaxation rates of the photophysics. As a consequence, the energy migration between the Dα and Dβ groups will be partially reversible. We refer to this case, as the partial donor–donor energy migration (PDDEM). The models of PPDEM presented can be used for analysing time-resolved fluorescence relaxation, as well as fluorescence depolarisation experiments. To explore the limitations of the PDDEM model, we have generated and re-analysed synthetic data that mimic time-correlated single photon counting (TCSPC) experiments. It was found that slow and fast rates of energy migration are most accurately recovered from the fluorescence relaxation and the depolarisation experiments, respectively. At comparable transfer and fluorescence rates, both kinds of experiments are equally useful. Real experiments on PDDEM were performed on an asymmetrically quenched bichromophoric molecule (1,32-dihydroxy-dotriacontane-bis-(Rhodamine 101) ester), that spans across the lipid bilayer of a vesicle. The depolarisation data were analysed by the PDDEM model and provide a distance between Rhodamine 101 groups, which agrees with independent studies

Place, publisher, year, edition, pages
2002. Vol. 58, no 5, 1087-1097 p.
Keyword [en]
Energy migration, Time-resolved fluorescence depolarisation, Intramolecular distances, Biomacromolecules
URN: urn:nbn:se:umu:diva-4159DOI: doi:10.1016/S1386-1425(01)00613-8OAI: diva2:143145
Available from: 2004-10-19 Created: 2004-10-19 Last updated: 2010-01-29Bibliographically approved
In thesis
1. Electronic Energy Transfer within Asymmetric Pairs of Fluorophores: Partial Donor-Donor Energy Migration (PDDEM)
Open this publication in new window or tab >>Electronic Energy Transfer within Asymmetric Pairs of Fluorophores: Partial Donor-Donor Energy Migration (PDDEM)
2004 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

A kinetic model of electronic energy migration within pairs of photophysically non-identical fluorophores has been developed. The model applies to fluorescent groups that exhibit different photophysical and spectral properties when attached to different positions in a macromolecule. The energy migration within such asymmetric pairs is partially reversible, which leads to the case of partial donor-donor energy migration (PDDEM). The model of PDDEM is an extension of the recently developed donor-donor energy migration model (DDEM, F. Bergström et al, PNAS 96 (1999) 12477), and applies to quantitative measurements of energy migration rates and distances within macromolecules. One important distinction from the DDEM model is that the distances can be obtained from fluorescence lifetime measurements. A model of fluorescence depolarisation in the presence of PDDEM is also presented.

To experimentally test the PDDEM approach, different model systems were studied. The model was applied to measure distances between rhodamine and fluorescein groups within on-purpose synthesised molecules that were solubilised in lipid bilayers. Moreover, distances were measured between BODIPY groups in mutant forms of the plasminogen activator inhibitor of type 2 (PAI-2). Measurements of both the fluorescence intensity decays and the time-resolved depolarisation were performed. The obtained distances were in good agreement with independent determinations.

Finally, the PDDEM within pairs of donors is considered, for which both donors exhibit a nonexponential fluorescence decay. In this case it turns out that the fluorescence relaxation of a coupled system contains distance information even if the photophysics of the donors is identical. It is also demonstrated that the choice of relaxation model has a negligible effect on the obtained distances. The latter conclusion holds also for the case of donor-acceptor energy transfer.

Place, publisher, year, edition, pages
Umeå: Kemi, 2004. 36 p.
Physical chemistry, fluorescence resonance energy transfer (FRET), donor-donor energy migration (DDEM), homotransfer, fluorescence relaxation, lifetimes, time-resolved fluorescence anisotropy, time-correlated single photon counting, distance measurements, protein structure, Fysikalisk kemi
National Category
Physical Chemistry
Research subject
Physical Chemistry
urn:nbn:se:umu:diva-338 (URN)91-7305-765-7 (ISBN)
Public defence
2004-11-19, KE 32, Kemihuset, Umeå University, Umeå, 13:00 (English)
Available from: 2004-10-19 Created: 2004-10-19 Last updated: 2010-08-05Bibliographically approved

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