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- Edman, Peter, et al.
(författare)
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Extended Förster theory of donor-donor energy migration in bifluorophoric macromolecules. Part II : Method for determining intramolecular distances with experimental validation using mono and bifluorophoric systems
- 2000
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Ingår i: Physical chemistry chemical physics. ; 2:12, s. 2795-2801
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Tidskriftsartikel (refereegranskat)abstract
- Recently an approximate theory was presented and applied for determining intramolecular distances in proteins. The rate of donor-donor energy migration (DDEM) is extracted and analysed from fluorescence depolarisation experiments by means of the DDEM model (Karolin et al., Biophys. J., 1998, 74, 11; Bergström et al., Proc. Natl. Acad. Sci., 1999, 96, 12477). Previously an extended Förster theory (EFT) was derived (Johansson et al., J. Chem. Phys., 1996, 105, 10896), which accounts for DDEM between reorienting molecules. For the first time, this rigorous theory is applied for analysing time-resolved fluorescence depolarisation data, accumulated by using the time-correlated single photon counting (TCSPC) technique. A simulation-deconvolution algorithm is presented which reduces the need of the DDEM model (Edman et al., Phys. Chem. Chem. Phys., 2000, 2, 1789), and other approximate theories (Edman et al., Mol. Phys., submitted). Two bifluorophoric systems were studied, namely; 1,32-dihydroxy-dotriacontane-bis(rhodamine) 101 ester solubilised in lipid vesicles, and bis(9-anthrylmethyl-phosphonate) bisteroid dissolved in propane-1,2-diol. The bis-rhodamine molecules span across lipid bilayers, so that the two rhodamine moieties of the molecule are localised on opposite sides of a bilayer. From the analyses of the fluorescence anisotropy, the donor-donor distances were determined to be 36.5 ± 1 and 21.0 ± 1.5 Å, for the membrane spanning molecule and the bisteroid, respectively. The results are in good agreement with independent studies.
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| 3. |
- Edman, Peter, et al.
(författare)
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On determining intramolecular distances from donor–donor energy migration (DDEM) within bifluorophoric macromolecules
- 2000
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Ingår i: PHYSICAL CHEMISTRY CHEMICAL PHYSICS. - : Royal Society of Chemistry (RSC). - 1463-9076 .- 1463-9084. ; 2, s. 1789-94
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Tidskriftsartikel (refereegranskat)abstract
- Recently a model based on donor–donor energy migration (DDEM) was developed for examining structure–function properties of biomacromolecules such as proteins (J. Chem. Soc., Faraday Trans., 1996, 92, 1563). Unlike the extended Förster theory (EFT; J. Chem. Phys., 1996, 105, 10896) the DDEM model is straightforward to apply in the analyses of fluorescence depolarisation experiments, as obtained by the time-correlated single photon counting (TCSPC) technique. In order to test the validity of the DDEM model, the EFT was used to create synthetic depolarisation data. These mimic true TCSPC experiments and cover a wide range of physical conditions, which are difficult to arrange for in real experiments with model systems. In particular, the relative rate of DDEM () and the rotation correlation time () of the donor molecules was examined. The DDEM rates obtained from the analyses were compared to the true rates. From these results the relative error of the intramolecular distances were calculated. For values 1<12<25, the DDEM model is slightly overestimating the distances. Typically, the distances determined with the DDEM model are overestimated by 5–10%.
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