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JungSookKang,JosephR.Lakowicz,GrzegorzPiszczek 대한약학회 2002 Archives of Pharmacal Research Vol.25 No.3
Fluorescent probes bound to DNA typically display nanosecond decay times and reveal only nanosecond motions. We extend the time range of measurable DNA dynamics using [Ru(bpy)2(dppz)]2+ (bpy=2,2'-bipyridine, dppz=dipyrido[3,2-a:2’,3’-c]phenazine) (RuBD) which displays a mean lifetime near 90 ns. To test the usefulness of RuBD as a probe for diffusive processes in calf thymus DNA, we compared the efficiencies of fluorescence resonance energy transfer (FRET) using three donors which display lifetimes near 5 ns for acridine orange (AO), 22 ns for ethidium bromide (EB) and 92 ns for RuBD, with nile blue (NB) as the acceptor. The Förster distances for AO-NB, EB-NB and RuBD-NB donor-acceptor pairs were 42.3, 52.3, and 30.6 Å, respectively. All three donors showed dramatic decreases in fluorescence intensities and more rapid intensity decays with increasing NB concentrations. The intensity decays of AO and EB in the presence of varying concentrations of NB were satisfactorily described by the one-dimensional FRET model without diffusion (Blumen and Manz, 1979). In the case of the long-lifetime donor RuBD, the experimental phase and modulation somewhat deviated from the recovered values computed from this model. The recovered NB concentrations and FRET efficiencies from the model were slightly larger than the expected values, however, the recovered and expected values did not show a significant difference. Thus, it is suggested that the lifetime of RuBD is too short to measure diffusive processes in calf thymus DNA.
(Jung Sook Kang),(Omoefe O. Abugo),(Joseph R. Lakowicz) 생화학분자생물학회 2002 BMB Reports Vol.35 No.4
[Ru(bpy)_2(dppz)]^2+ (bpy = 2,2`-bipyridine, dppz = dipyrido-[3,2-a:2`,3`-c]phenazine) (RuBD), a long-lifetime metal-ligand complex, displays favorable photophysical properties. These include long lifetime, polarized emission, but no significant fluorescence from the complex that is not bound to DNA. To show the usefulness of this luminophore (RuBD) for probing the bending and torsional dynamics of nucleic acids, its intensity and anisotropy decays when intercalated into supercoiled and relaxed pTZ18U plasmids were examined using frequency-domain fluorometry with a blue light-emitting diode (LED) as the modulated light source. The mean lifetimes for the supercoiled plasmids (<τ> = 148 ns) were somewhat shorter than those for the relaxed plasmids (<τ> = 160 ns). This suggests that the relaxed plasmids were shielded more efficiently from water. The anisotropy decay data also showed somewhat shorter slow rotational correlation times for supercoiled plasmids (288 ns) than for the relaxed plasmids (355 ns). The presence of two rotational correlation times suggests that RuBD reveals both the bending and torsional motions of the plasmids. These results indicate that RuBD can be useful for studying both the bending and torsional dynamics of nucleic acids.
( Jung Sook Kang ),( Byeng Wha Son ),( Hong Dae Choi ),( Ji Hye Yoon ),( Woo Sung Son ) 생화학분자생물학회 2005 BMB Reports Vol.38 No.1
We extended the measurable time scale of DNA dynamics to microsecond using [Ru(phen)₂(dppz)^(2+) (phen = 1,10-phenanthroline, dppz = dipyrido[3,2-a:2`,3`-c]phenazine) (RuPD), which displays a mean lifetime near 500 ns. To evaluate the usefulness of this luminophore (RuPD) for probing nucleic acid dynamics, its intensity and anisotropy decays when intercalated into supercoiled and linear pBluescript (pBS) Ⅱ SK(+) phagemids were examined using frequency-domain fluorometry with a blue light-emitting diode (LED) as the modulated light source. The mean lifetime for the supercoiled phagemids (<τ> = 489.7 ns) was somewhat shorter than that for the linear phagemids (<τ> = 506.4 ns), suggesting a more efficient shielding from water by the linear phagemids. The anisotropy decay data also showed somewhat shorter slow rotational correlation times for supercoiled phagemids (997.2 ns) than for the linear phagemids (1175.6 ns). The slow and fast rotational correlation times appear to be consistent with the bending and torsional motions of the phagemids, respectively. These results indicate that RuPD can have applications in studies of both bending and torsional dynamics of nucleic acids.