CD Study of the G-Quadruplex Conformation
Circular Dichroic (CD) spectroscopy is one of the most frequently used methods for guanine quadruplex studies and in general for studies of conformational properties of nucleic acids. The reason is its high sensitivity to even slight changes in mutual ori
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troduction Circular Dichroism is a phenomenon that occurs when linearly polarized light passes through absorbing optically active media. Optically active materials (i.e., substances containing chiral molecules) rotate the plane of linearly polarized light due to different refractive indices of its left and right circularly polarized components, thus giving rise to optical rotation. If a studied chiral substance absorbs light, the two components are absorbed differently, resulting in the change of the originally linearly polarized light into elliptically polarized light. The difference in the absorption of the left and right circularly polarized light is called circular dichroism (CD). It is expressed by Δε ¼ εL ‑ εR, in units of [M1 cm1], where εL and εR are molar absorption coefficients of the two light components. Other quantity, which measures this phenomenon, is called ellipticity, Θ. The ellipticity corresponds to the angle whose tangent is the ratio between the minor and major axes of the
Danzhou Yang and Clement Lin (eds.), G-Quadruplex Nucleic Acids: Methods and Protocols, Methods in Molecular Biology, vol. 2035, https://doi.org/10.1007/978-1-4939-9666-7_2, © Springer Science+Business Media, LLC, part of Springer Nature 2019
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resulting elliptically polarized light. Molar ellipticity is expressed in units of [deg cm2 dmol1]. There are two conditions for an emergence of a CD signal: a sample of interest must be chiral and must absorb light. In the case of nucleic acids (NA), the chirality is fulfilled by sugar, deoxyribose or ribose containing asymmetric carbon C10 , and the absorption is accomplished by nucleic acid bases. To structural studies of nucleic acids, CD spectra are measured in the ultraviolet region of light (wavelengths ranging from 200–330 nm) corresponding to electronic transitions. Consequently, mononucleosides already provide CD signal. However, the main source of nucleic acid chirality arises from their secondary structure, i.e., from the folding of absorbing bases into asymmetric helical arrangements. CD thus reflects even slight changes in the mutual orientations of absorbing units. Due to its unique sensitivity to the conformation of nucleic acids, CD spectroscopy has become frequently used method in the field and it has been participating, very often as a pioneering method, in all basic findings on DNA conformational flexibility [1]. DNA can adopt, depending on its primary structure, various arrangements that are distinctly different from the classical Watson–Crick model. These noncanonical structures may differ in base pairing, the sense of the helix winding, and in the number and mutual orientation of oligonucleotide chains in a molecule. The individual structures provide characteristic CD spectra. CD spectroscopy was, e.g., the first method to discover the left-handed form of Z-DNA [2], several years before its existence was demonstrated in the crystal [3]. Nowadays, CD is frequently used in the study of G-quadruplexes, which are a hot topic of curre
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