| 1. |
- Engman, Cecilia, 1974, et al.
(författare)
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DNA adopts normal B-form upon incorporation of highly fluorescent DNA base analogue tC: NMR structure and UV-Vis spectroscopy characterization
- 2004
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Ingår i: Nucleic Acids Research. - : Oxford University Press (OUP). - 1362-4962 .- 0305-1048. ; 32:17, s. 5087-95
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Tidskriftsartikel (refereegranskat)abstract
- The influence of the highly fluorescent tricyclic cytosine base analogue (tC) on duplex DNA conformation is investigated. The duplex properties are characterized by absorbance and circular dichroism (CD) for all combinations of neighbouring bases to tC, and an NMR structure is determined for one tC-containing sequence. For the oligonucleotides with one tC incorporated instead of cytosine, the melting temperature is increased on average by 2.7 degrees C above that for the unmodified ones. CD spectra are practically identical for modified and unmodified sequences, indicating an unperturbed B-DNA conformation. The NMR structure determination of the self-complementary sequence 5'-CTC(tC)ACGTGGAG shows a DNA conformation consistent with B-form for the whole duplex. The root-mean-square distance for the nucleotides of the eight central base pairs between the 10 structures with lowest CYANA target functions and a mean structure is 0.45 +/- 0.17 A. The NMR data confirm correct base pairing for tC by the observation of both intrastrand and interstrand imino proton NOEs. Altogether, this suggests that tC works well as a cytosine analogue, i.e. it is situated in the base stack, forming hydrogen bonds with G in the complementary strand, without distorting the DNA backbone conformation. This first example of an artificial, highly fluorescent DNA base that does not perturb the DNA conformation could have valuable applications for the study of the structure and dynamics of nucleic acid systems.
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| 2. |
- Olofsson, Johan, 1974, et al.
(författare)
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Effects of methyl substitution on radiative and solvent quenching rate constants of [Ru(phen)(2)dppz](2+) in polyol solvents and bound to DNA
- 2004
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Ingår i: Journal of the American Chemical Society. - : American Chemical Society (ACS). - 1520-5126 .- 0002-7863. ; 126:47, s. 15458-15465
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Tidskriftsartikel (refereegranskat)abstract
- Methyl substituents on the distant benzene ring of the dppz ligand in the "light switch" complex [Ru(phen)(2)dppz](2+) have profound effects on the photophysics of the complexes in water as well as in the polyol solvents ethylene glycol, glycerol, and 1,2- and 1,3-propanediol. Whereas 11,12-dimethyl substitution decreases the rate of quenching by diminishing hydrogen bonding by solvent, the 10-methyl substituent in addition also decreases both the radiative and the nonradiative rate constant for decay to the ground state of the non-hydrogen-bonded excited state species. For both the 10-methyl and the 11,12-dimethyl derivatives, the effect of methyl substitution on the equilibrium of solvent hydrogen bonding to the excited state is due to changes in the entropy terms, rather than in the enthalpy; indicating that the effect is a steric perturbation of the solvent cage around the molecule. When intercalated into DNA, the effects of methyl substitution is smaller than those in polyol solvent or water, suggesting that the water molecules that quench the excited state by hydrogen bonding to the phenazine aza nitrogens mainly access them from the same groove as in which the Ru(II) ion resides. Since the Delta-enantiomer of [Ru(phen)(2)10-methyl-dppz](2+) has an absolute quantum yield of up to 0.23 when bound to DNA, a value 7000 times higher than in pure water solution, it is promising as a new luminescent DNA probe.
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| 3. |
- Westerlund, Fredrik, 1978, et al.
(författare)
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Micelle-sequestered dissociation of cationic DNA-intercalated drugs: Unexpected surfactant-induced rate enhancement
- 2003
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Ingår i: Journal of the American Chemical Society. - : American Chemical Society (ACS). - 1520-5126 .- 0002-7863. ; 125:13, s. 3773-3779
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Tidskriftsartikel (refereegranskat)abstract
- Detergent sequestration using micelles as a hydrophobic sink for dissociated drug molecules is an established technique for determination of dissociation rates. The anionic surfactant molecules are generally assumed not to interact with the anionic DNA and thereby not to affect the rate of dissociation. By contrast, we here demonstrate that the surfactant molecules sodium dodecyl sulfate (SDS), sodium decyl sulfate, and sodium octyl sulfate all induce substantial rate enhancements of the dissociation of intercalators from DNA. Four different cationic DNA intercalators are studied with respect to surfactant-induced dissociation. Except for the smallest intercalator, ethidium, the dissociation rate constants increase monotonically with surfactant concentration both below cmc and (more strongly) above cmc, much more than expected from electrostatic effects of increased counterion concentration. The rate enhancement, most pronounced for the bulky, multicationic, hydrophobic DNA ligands in this study, indicates a reduction of the activation energy for the ligand to pass out from a deeply penetrating intercalation site of DNA. The discovery that surfactants enhance the rate of dissociation of cationic DNA-intercalators implies that rate constants previously determined by micelle-sequestered dissociation may have been overestimated. As an alternative, more reliable method, we suggest instead the addition of excess of dummy DNA as an absorbent for dissociated ligand.
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| 4. |
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| 5. |
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| 6. |
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| 7. |
- Wilhelmsson, Marcus, 1974, et al.
(författare)
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Genetic screening using the colour change of a PNA-DNA hybrid-binding cyanine dye
- 2002
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Ingår i: Nucleic Acids Research. - : Oxford University Press (OUP). - 0305-1048 .- 1362-4962. ; 30:2
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Tidskriftsartikel (refereegranskat)abstract
- As the relationship between human genes and various malfunctions and diseases becomes revealed at an ever-increasing pace, the need arises for the development of rapid genetic screening methods for diagnostic purposes. Genetic diseases show great diversity. Some are caused by a few characteristic localised mutations, while others arise from a large number of variations. Hence, It. is unlikely that. a single, general diagnostic method that applies to all cases will ever exist. Instead, a combination of methods is frequently applied. Here we propose the use of a dramatic colour change that a cyanine ye, 3,3'-diethylthiadicarbocyanine, displays upon binding to DNA-PNA duplexes. This method could become an inexpensive, fast and simple genetic screening test by visual inspection, with no need for complicated equipment. Our results demonstrate that this diagnostic method may be sufficiently sensitive to discriminate between even a fully complementary and a single mutation DNA sequence.
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| 8. |
- Wilhelmsson, Marcus, 1974, et al.
(författare)
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Meso stereoisomer as a probe of enantioselective threading intercalation of semirigid ruthenium complex [µ-(11,11'-bidppz)(phen)4Ru2]4
- 2003
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Ingår i: Journal of Physical Chemistry B. - : American Chemical Society (ACS). - 1520-5207 .- 1520-6106. ; 107:42, s. 11784-11793
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Tidskriftsartikel (refereegranskat)abstract
- Upon interaction with calf thymus DNA the Lambda,Lambda-enantiomer of the semirigid binuclear ruthenium complex [mu-(11, 11'-bidppz)(phen)(4)Ru-2](4+) has previously been shown to reorganize from an initial groove bound geometry into an intercalative binding mode, threading one of its bulky Ru(phen)(2) moieties through the core of the DNA. We have now found that all three stereoisomers, Delta,Delta; Lambda,Lambda; and Delta,Lambda (meso), are intercalated in their final modes of binding to calf thymus DNA, poly(dA-dT)(2), poly(dG-dC)(2), as well as poly(dI-dC)(2) indicated by linear dichroism, circular dichroism, and luminescence. For all three stereoisomers, studied in detail with poly(dA-dT)(2), the bridging bidppz ligand is intercalated in anti conformation, leaving one Ru(phen)(2) Moiety in each groove. This final binding geometry is characterized by a distinct clockwise roll of the Ru(phen)(2) moiety in the minor groove, similar to the roll earlier observed for the dppz ligand in [Ru(phen)(2)dppz](2+). Using the meso stereoisomer as an enantioselective probe, it is shown that the Lambda moiety prefers to insert itself deeply into the minor groove while the Delta moiety, in the major groove, is somewhat displaced from the center of the DNA helix. The preceding, metastable bound geometries are concluded to be in the major groove for calf thymus DNA, poly(dG-dC)(2), and poly(dI-dC)(2), with the Delta,Delta form displaying an angle of the bidppz bridge relative the DNA helix axis of about 50degrees, whereas the corresponding angles for the meso- and Lambda,Lambda-forms in calf thymus DNA are around 65degrees, suggesting an orientation in the groove more parallel to the bases. By contrast, in poly(dA-dT)(2) none of the stereoisomers exhibits any distinguishable initial groove binding mode, but all seem to bind by threading intercalation directly.
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| 9. |
- Wilhelmsson, Marcus, 1974, et al.
(författare)
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Photophysical Characterization of Fluorescent DNA Base Analogue, tC
- 2003
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Ingår i: Journal of Physical Chemistry B. - : American Chemical Society (ACS). - 1520-5207 .- 1520-6106. ; 107:34, s. 9094-9101
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Tidskriftsartikel (refereegranskat)abstract
- The novel fluorescent DNA cytosine base analogue 1,3-diaza-2-oxophenothiazine, tC, has previously been shown to have a remarkably preserved high quantum yield upon incorporation into a single strand of peptide nucleic acid, PNA, as well as when the latter is hybridized with a complementary DNA to form a PNA-DNA duplex. Here, we investigate fundamental photophysical properties of tC. Using fluorescence anisotropy, stretched film linear dichroism, and quantum chemical calculations, the transition moment polarizations of the lowest lying electronic states are determined. The neutral, base-pairing form of tC, having a fluorescence quantum yield of 0.2, is found to be the totally predominant species in a wide pH interval, 4-12. We show that the absorption band of tC at lowest energy, centered at 26 700 cm(-1) and well separated from the nucleobase absorption, is due to a single electronic transition polarized approximately at 35degrees from the long axis of the molecule. The 2-deoxyribonucleoside of 1,3-diaza-2-oxophenothiazine, synthesized for further incorporation into DNA, was found to display a fluorescence quantum yield nearly the same as in the form of tC that was incorporated into PNA, confirming the notion of the tC nucleoside being a probe with very promising fluorescence properties essentially invariant of environment, also upon incorporation into a DNA strand and upon hybridization.
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| 10. |
- Wilhelmsson, Marcus, 1974
(författare)
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Recognition and Visualization of Nucleic Acids
- 2003
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The work presented in this Thesis focuses on the interaction between nucleic acids and small molecules, as well as binding or incorporation of reporter chromophores into nucleic acids. The results could help in studying and understanding processes that are fundamental to describe mechanisms behind a genetic disorder. For example, the molecules investigated could be used for studying the mechanism of interaction between DNA and other molecules, such as proteins or drugs, and also give information about intrinsic properties of DNA during, for example, DNA-protein interaction. In Paper I, a dramatic color change of a cyanine dye, 3,3'-diethylthiadicarbocyanine, is used in the development of a fast and straightforward low-cost assay for genetic screening. The aggregation of several dyes within the minor groove of the double helix, responsible for the color change of the dye upon binding to PNA-DNA duplexes, is also in itself a very interesting observation, giving information about the molecular dimensions of the binding site. Paper II and III reports on thermodynamic and kinetic DNA-binding properties of the dimeric compound, [μ(11,11'-bidppz)(phen)4Ru2]4+. Flow linear dichroism and luminescence studies show that the ruthenium complex extremely slowly rearranges from an initial groove-bound to a final threaded intercalated geometry. The slowness of this rearrangement of DNA-binding modes, due to the requirement of opening of at least on base pair, is to the best of our knowledge, unprecedented. In Paper IV, the SDS-sequestering method for measuring dissociation of cationic drugs from DNA is reported to strongly overestimate the rate of dissociation of bulky multicationic intercalators. An explanation, where the surfactant molecules are involved in a reduction of the activation barrier for the cationic intercalators to leave DNA, is presented as well as an alternative, more reliable method for determining dissociation rates based on use of dummy DNA as an absorbent for the dissociated species. In Paper V and VI, a novel fluorescent base analogue, tC, is presented. Its high fluorescence quantum yield (0.2) is preserved upon incorporation into nucleic acid and subsequent hybridization to a complementary DNA strand. The minimal influence of environment on the photophysical properties of tC and its firm stacking between the bases, makes it very promising as a probe of intra- and inter-molecular distance estimations in nucleic acid systems.
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