Polarity Sensing by Fluorescent Oligonucleotides:  First Demonstration of Sequence-Dependent Microenvironmental Changes in the DNA Major Groove

Abstract

DNA duplex recognition by macromolecules (proteins, enzymes) and small molecules (drugs, metal complexes) occurs in the major or minor grooves of DNA via hydrogen bonding and electrostatic or hydrophobic interactions, whose strengths depend on the medium. It is therefore important to understand the local environments of the major/minor grooves of DNA. It is known that small molecules bind in the minor groove that have nonpolar character (ε = 22) and are organic-like. By employing fluorescent oligonucleotides, we demonstrated recently that the major groove of DNA is more polar than the minor groove (ε = 55) and the DNA double helix has different polarity in its two grooves. Our experimentally measured values were recently validated independently by two theoretical groups, employing calculations by two different methodologies. In this paper, a hitherto unknown property of DNA double helixsequence dependent local polarity or microenvironmentis experimentally demonstrated. This is shown by fluorescence experiments using oligonucleotides containing the modified base 5-amino-dU in place of dT and the polarity varied between ε ≈ 40 and 60, in different sequences. This is the first experimental demonstration of sequence-dependent microenvironmental effects in DNA in solution, although structural effects are well-known by X-ray data. The technique used has a good potential to investigate differing microenvoronmental effects in various secondary structures, DNA polymorphs, and chemically modified DNA and their hybrids.

Fluorescent oligonucleotides containing a novel perylene 2′-amino-α-L-LNA monomer: Synthesis and analytical potential

“…60 nm red shifted relative to the corresponding probes containing pyrene 2′-amino-α-L-LNA 27 . Upon duplex formation the fluorescence maximum of ON5 shifts 9-10 nm to shorter wavelengths which points on direction of the two fluorophores to the less polar microenvironment compared to the aqueous buffer 25,40,41 . Emission maxima of the dual-probe system ON6+ON7 on the contrary are only slightly Modified LNA/DNA mixmer oligonucleotides prepared in this study: ON5: 5′-d(CA L C CAA CT*T* CT L T CCA C L A), ON6: 5′-d(CA L C CAA CT*), ON7: 3′-d(AC L A CCT T L CT*), where A L , T L and C L are adenine-9-yl, thymin-1-yl and 5-methylcytosin-1-yl LNA monomers, respectively.…”

Polarity of Major Grooves Explored by Using an Isosteric Emissive Nucleoside

“…[5,6] The polarity of DNA grooves has been experimentally interrogated by using environmentally sensitive fluorescent probes. [7][8][9][10][11][12][13][14][15][16] This approach is informative, but bears several predicaments: a) any probe placed within the cavity to be assessed inherently modifies the molecular architecture of the native environment and therefore taints the readout; b) most studies have utilized dielectric constant (e, relative permittivity)-a parameter that defines bulk solvent property and not an anisotropic medium as its measure; c) relatively large fluorophores (e.g., dansyl), which are frequently connected by flexible linkers, have been employed; these possibly populate multiple conformers each of which senses a different microenvironment. [17] Collectively, these challenges are responsible, at least in part, for the dramatically different estimates so far reported for the dielectric constant of the major groove in nucleic acids, which range from about 40 to 70.…”

Monte Carlo simulations for a fluctuating sphere labeled on a flexible polymer chain in good solvents

“…In other words, the fluctuating sphere is coupled with the geometry of the grafted segment when a grafted segment is present. These findings provide physical insights regarding the confinement effects induced by a flex- ible chain polymer, and may be useful in reasoning the role of a DNA molecule on drug-DNA binding 8,9 or a polymer molecule on the labeled probe.…”

“…7 An important system similar to the probe labeling problem is the drug-DNA binding in biomedical research, in which bio-active molecules are bound to polymeric molecules. 8,9 Further, not only does a small molecule bind with polymeric molecules, but also a chain segment, such as the protein molecule in the enzyme-RNApolymerase. 10 Recently, a novel experimental method has been developed to facilitate the crystallization of RNA molecules via labeling a bulky protein segment onto a long chain RNA.…”

“…These findings provide physical insights regarding the confinement effect induced by a flexible chain polymer, and may be useful in reasoning the role of the long chain DNA molecule on drug-DNA binding. 8,9 Also, the behavior of the vibrational spectroscopy of a labeled chromophore subject to a grafted bound segment is summarized in the Appendix to illustrate the confinement effect on a labeled molecule.…”