Pressure Perturbation Studies of Noncanonical Viral Nucleic Acid Structures

SIMPLE SUMMARY: It is well known that nucleic acids adopt a double-helical structure that is essential in the storage and replication of genetic code. However, noncanonical structures of DNA are less explored. The most important of these structures are the G-quadruplexes, which are formed by guanine-rich sequences of the genome. These G-quadruplexes were found in several crucial positions of the genome; they regulate important processes such as cell proliferation and death. Here we are investigating the quadruplex structures appearing in the genome of the hepatitis B, whose infection is among the ten leading causes of death. Our unique approach—the high-pressure technique—allowed us to characterize the stability of these quadruplexes in wide temperature and pressure ranges. Pressure experiments gave us the volume changes occurring at the unfolding, e.g., the embedded volume of the folded structure. Volumetric parameters are especially important because the space available for molecules is quite limited in the crowded environment of the cell. We also investigated how the stability of the viral quadruplexes can be increased by the binding of TMPyP4, a special ligand developed for cancer therapy. ABSTRACT: G-quadruplexes are noncanonical structures formed by guanine-rich sequences of the genome. They are found in crucial loci of the human genome, they take part in the regulation of important processes like cell proliferation and cell death. Much less is known about the subjects of this work, the viral G-quadruplexes. We have chosen three potentially G-quadruplex-forming sequences of hepatitis B. We measured the stability and the thermodynamic parameters of these quadruplexes. We also investigated the potential stabilization of these G-quadruplexes by binding a special ligand that was originally developed for cancer therapy. Fluorescence and infrared spectroscopic measurements were performed over wide temperature and pressure ranges. Our experiments indicate the small unfolding volume change of all three oligos. We found a difference between the unfolding of the 2-quartet and the 3-quartet G-quadruplexes. All three G-quadruplexes were stabilized by TMPyP4, which is a cationic porphyrin developed for stabilizing the human telomere.