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We have found that the efficiency of fluorescence resonance energy transfer between Cy3 and Cy5 terminally attached to the 5 ends of a DNA duplex is significantly affected by the relative orientation of the two fluorophores. The cyanine fluorophores are predominantly stacked on the ends of the helix in the manner of an additional base pair, and thus their relative orientation depends on the length of the helix. Observed fluorescence resonance energy transfer (FRET) efficiency depends on the length of the helix, as well as its helical periodicity. By changing the helical geometry from B form double-stranded DNA to A form hybrid RNA/DNA, a marked phase shift occurs in the modulation of FRET efficiency with helix length. Both curves are well explained by the standard geometry of B and A form helices. The observed modulation for both polymers is less than that calculated for a fully rigid attachment of the fluorophores. However, a model involving lateral mobility of the fluorophores on the ends of the helix explains the observed experimental data. This has been further modified to take account of a minor fraction of unstacked fluorophore observed by fluorescent lifetime measurements. Our data unequivocally establish that Fö rster transfer obeys the orientation dependence as expected for a dipole-dipole interaction.cyanine fluorophores ͉ FRET ͉ kappa squared ͉ single-molecule FRET F luorescence resonance energy transfer (FRET) has become widely used to report on distances over the macromolecular scale in biology (1), reviewed in refs. 2-4. The method is highly sensitive, and consequently has been widely exploited in singlemolecule experiments in biological systems. Energy transfer results from dipolar coupling between the transition moments of two fluorophores, and the efficiency of the process (E FRET ) depends on the separation between the donor and acceptor fluorophores, raised to the sixth power. Although such data are frequently interpreted on the assumption of a simple relationship between E FRET and distance, E FRET should also depend on the relative orientation of the transition dipole vectors.The orientation dependence is likely to be most significant where the fluorophores are constrained (5-9). This has been demonstrated experimentally by using a fluorophore that was terminally affixed to duplex DNA by two points of covalent attachment (10), thereby seriously constraining its motion. This situation is not typical of most FRET studies involving nucleic acids. Fluorophores are normally tethered by a single point of attachment, and in theory would be significantly less constrained. But if the fluorophores adopt a rigid manner of attachment to the helix, an orientational dependence could be observed.Cy3 and Cy5 are a commonly used fluorophore pair, especially in single-molecule experiments. Our earlier NMR studies have shown that when these are attached to the 5Ј termini of duplex DNA via a 3-carbon linker to the 5Ј-phosphate they are predominantly stacked onto the ends of the helix in the manner of an additio...

Ultrafast solvent dynamics of room-temperature ionic liquids have been investigated by optical heterodyne-detected Raman-induced Kerr-effect spectroscopy ~OHD-RIKES! by studying the effects of cation and anion substitution on the low frequency librational modes. The spectra of two series of imidazolium salts are presented. The first series is based on the 1-butyl-3-methylimidazolium salts @bmim#1 containing the anions trifluoromethanesulfate @TfO#2, bis~trifluoromethanesulfonyl!imide @Tf2N#2, and hexafluorophosphate @PF6#2. The second series is based on @Tf2N#2 salts containing the three cations 1-butyl-2,3-dimethylimidazolium @bmmim#1, 1-methyl-3-octylimidazolium @omim#1, and @bmim#1. It is found in all five samples that the signal is due to libration of the imidazolium ring at three frequencies around 30, 65, and 100 cm21 corresponding to three local configurations of the anion with respect to the cation

The low-frequency (1-200 cm(-1)) vibrational spectra of peptides and proteins in solution have been investigated with ultrafast optical heterodyne-detected Raman-induced Kerr-effect spectroscopy (OHD-RIKES). Spectra have been obtained for di-L-alanine (ALA(2)) and the alpha-helical peptide poly-L-alanine (PLA) in dichloroacetic acid solution. The poly-L-alanine spectrum shows extra amplitude compared to the di-L-alanine spectrum, which can be explained by the secondary structure of the former. The globular proteins lysozyme, alpha-lactalbumin, pepsin, and beta-lactoglobulin in aqueous solution have been studied to determine the possible influence of secondary or tertiary structure on the low-frequency spectra. The spectra of the globular proteins have been analyzed in terms of three nondiffusive Brownian oscillators. The lowest frequency oscillator corresponds to the so-called Boson peak observed in inelastic neutron scattering (INS). The remaining two oscillators are not observed in inelastic neutron scattering, do therefore not involve significant motion of hydrogen atoms, and may be associated with delocalized backbone torsions.

The low-frequency spectra of peptides and proteins in solution have been investigated with optical heterodyne-detected Raman-induced Kerr-effect spectroscopy. Spectra were obtained for di-l-alanine ALA(2) and poly-l-alanine (PLA) in dichloroacetic acid solution. The conformational dependence of those spectra at low frequency has been analyzed. ALA(2) displays a band centered at 50 cm-1, whereas the alpha-helical PLA shows two shoulders at 60 and 140 cm-1. The similarity of the spectral features observed in PLA to those in water can be explained by analogous acoustic translational modes in the hydrogen network of the PLA alpha-helix. The mostly alpha-helical protein lysozyme in aqueous solution has also been investigated and showed significantly more structure with modes at 10, 35, 73, 106, and 164 cm-1.

Abstract. Electrical and optical properties of binary inhomogeneous media are currently modelled by a random network of metallic bonds (conductance σ 0 , concentration p) and dielectric bonds (conductance σ 1 , concentration 1 − p). The macroscopic conductivity of this model is analytic in the complex plane of the dimensionless ratio h = σ 1 /σ 0 of the conductances of both phases, cut along the negative real axis. This cut originates in the accumulation of the resonances of clusters with any size and shape. We demonstrate that the dielectric response of an isolated cluster, or a finite set of clusters, is characterised by a finite spectrum of resonances, occurring at well-defined negative real values of h, and we define the cross-section which gives a measure of the strength of each resonance. These resonances show up as narrow peaks with Lorentzian line shapes, e.g. in the weakdissipation regime of the RL − C model. The resonance frequencies and the corresponding cross-sections only depend on the underlying lattice, on the geometry of the clusters, and on their relative positions. Our approach allows an exact determination of these characteristics. It is applied to several examples of clusters drawn on the square lattice. Scaling laws are derived analytically, and checked numerically, for the resonance spectra of linear clusters, of lattice animals, and of several examples of self-similar fractals.

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