Projects / Programmes
Pre-folded structures of DNA G-quadruplexes
| Code |
Science |
Field |
Subfield |
| 1.04.02 |
Natural sciences and mathematics |
Chemistry |
Structural chemistry |
| Code |
Science |
Field |
| P003 |
Natural sciences and mathematics |
Chemistry |
| Code |
Science |
Field |
| 1.04 |
Natural Sciences |
Chemical sciences |
DNA, quadruplex, intermediates, NMR, folding
Researchers (8)
Organisations (2)
Abstract
In addition to a well-known right-handed B form double helix, DNA molecules can form higher order structures such as G-quadruplexes. Although DNA is most widely known to store and pass along genetic information, the discovery of G-quadruplex structures has illuminated a new role of DNA in biology. G-quadruplexes are very stable secondary structures formed by guanine-rich DNA. Those sequences have been found in regions with biological significance, such as human telomeres and oncogene-promoter regions. Therefore, the formation of G-quadruplexes is closely related to human diseases. In order to understand how cells work, it is crucial to identify their components and characterize their interactions. The function of a molecule is tributary of its three-dimensional structure. G-quadruplexes are structurally very polymorphic. G-quartet, the main building block of G-quadruplexes, requires the presence of cations for its formation in order to reduce repulsion between carbonyl oxygen atoms on C6 of guanine. Cations are located in the central channel of G-quadruplex core and they can move between binding sites and bulk solution. The direction of cation movements is governed by structural features. The development of small molecules that can bind and thereby help in the formation and stabilize G-quadruplex structures motivate in search of anticancer and antiviral drugs. G-quadruplexes can also be used as building blocks in functional materials and biotechnology due to their structural diversity and high temperature stability. In contrast to the double-stranded DNA structure, in the case of quadruplexes there are many things that need to be understood. Recent studies utilizing NMR spectroscopy and other methods made in our laboratory have shown that hydrodynamic dimensions of model G-rich oligonucleotides without added salt do not correspond to an extended cylindrical shape of a linearized oligomer. Therefore in the absence of cations these G-rich sequences can self-assemble into specific pre-organized structures that could fold very rapidly and not by chance into a large number of structurally diverse G-quadruplex structures with mechanisms of varying complexity. G-quadruplex formation by telomeric repeats and gene regulatory G-rich regions could simply go through populating various intermediate states, which are generally unstable and hard to detect. Till now a lot of research has been done in the field of structural determination of G-quadruplex structures formed by different oligonucleotide sequences derived from telomeric and promoter regions of genes of humans and other organisms. However, to the best of our knowledge only two publications were dealing with the determination of detailed structural features of guanine rich DNA oligonucleotide before G-quadruplexes are formed.
The aim of our research work is to deepen our knowledge regarding mechanisms of G-quadruplex formation and their new structural features with emphasis on their pre-folded structures. The current project will make use of high resolution NMR spectroscopy in order to ascertain the nature of hydrogen bonds between nucleotides comprising pre-folded or G-quadruplex structures and for their 3D structure determination. For additional characterization of pre-folded and G-quadruplex structures native gel electrophoresis, thermal differential spectra (TDS), CD spectroscopy, UV spectroscopy and differential scattering calorimetry (DSC) will be utilized as well.
Significance for science
We expect that the results anticipated from the current proposal will contribute to the understanding of various structural features of DNA quadruplex structures as well as their pre-folded forms. Till now a lot of research has been done in the field of structural determination of G-quadruplex structures formed by different oligonucleotide sequences derived from telomeric and promoter regions of genes of humans and other organisms. However, to the best of our knowledge only two publications were dealing with the determination of detailed structural features of guanine rich DNA oligonucleotide before G-quadruplexes are formed. Such knowledge can also explain the structural polymorphism of G-quadruplex structures as well as kinetics of folding. Therefore, these kinds of studies where intermediate states can be characterized structurally, thermodynamically and kinetically provide an important step in the quest to elucidate general principles by which G-quadruplexes adopt their native folds. Understanding of G-quadruplex formation will contribute to more accurate planning of new G-quadruplex ligands, which could specifically bind only to a precise structure as well as in the design of nano machines on the basis of the oligonucleotides. Knowing of pre-folded structures will allow their targeting already at the stage before the formation of G-quadruplex, which could play a critical role in biological function and open up additional channel in regulating gene expression by manipulating the pre-folded structures prior to G-quadruplex transition. We will also contribute to the development of experimental methods for DNA structure elucidation.
Significance for the country
The research of correlations between the structure and function is highly relevant for pharmaceutical industry. Our studies may contribute to help in development of new areas of research and science in Slovenia, in particular in the understanding of various structural features of DNA quadruplex structures as well as their pre-folded forms. All of this could be used as targets for developing of new drugs as well as in the design of nano machines on the basis of the oligonucleotides. New drugs, in turn, will generate investments from pharmaceutical companies and may ultimately lead to the creation of new jobs in Slovenia.
Most important scientific results
Final report
Most important socioeconomically and culturally relevant results
Interim report,
final report
