Projects / Programmes
Studies of 3D structures of promotor sequences and their interactions with ligands
| Code |
Science |
Field |
Subfield |
| 1.04.02 |
Natural sciences and mathematics |
Chemistry |
Structural chemistry |
| Code |
Science |
Field |
| B002 |
Biomedical sciences |
Biophysics |
| Code |
Science |
Field |
| 1.04 |
Natural Sciences |
Chemical sciences |
DNA, quadruplex, NMR spectroscopy, three-dimensional structure
Researchers (8)
Organisations (3)
Abstract
Nucleic acids adopt a great number of topologies including multi-stranded helical structures in order to perform different biological functions from transport of genetic information to catalysis and regulation. Within the framework of the current proposal we propose to study structural features of guanine rich genomic DNA sequences, which are expected to form G-quadruplex structures. Bioinformatic studies have revealed more than 376.000 distinct sites in the human genome with potential to form G-quadruplex structures. We will primarily focus on promoter regions of genes. G-quadruplex structures formed by promoters have been in comparison to telomeric regions addressed in only few studies and their structural features are poorly understood, which raises the need to determine their structures. In many promoter regions of the human genome, including the important proto-oncogenes such as c-myc, VEGF, HIF-1alpha, ret, KRAS, bcl-2, c-kit, PDGF-A and c-myb guanine-rich regions were observed which may form G-quadruplex structure. Numerous findings support the hypothesis that formation and unfolding of specific G-quadruplex structures may be involved in regulation of gene expression. The basic building block of G-quadruplex is G-quartet in which four guanine bases are held together by eight Hoogsteen hydrogen bonds in a co-planar arrangement. G-quadruplexes can be formed from a single, two or even four oligonucleotides. Past studies have shown that G-quadruplexes are highly polymorphic, which has been associated with specifics of oligonucleotide sequences, as well as with conditions in solution. Connecting loops, which can span diagonal or edge of the outer G-quartet or can adopt a double chain reversal orientation along the side of G-quadruplex core, play very important role in stabilization of G-quadruplex structures. We will devote special attention to the nature and sequence requirements of loops since their structure is very important in terms of interaction between DNA molecule and ligands (potential drugs). The primary research tool will be high resolution NMR spectroscopy. We will complement the results by UV, CD and other (spectroscopic) methods. In addition to characterizing 3D structures with atomic resolution we will also study dynamics of individual parts of the structure, especially loop regions. Structural information will be used in the design of new small molecule ligands with potential to regulate expression of a gene through (de)stabilization of G-quadruplex structure.
Significance for science
The results expected from our research work should contribute to expansion of knowledge on folding of promoter sequences into G-quadruplex structures and on their dynamic properties. Up till now, very little is known about G-quadruplex structures adopted by G-rich regions found in promoters. Such knowledge is important for understanding the regulation of gene expression and can be of help in design of novel organic molecules that could specifically stabilize certain G-quadruplex structure. This will help against various forms of cancer and many viral diseases. Alternatively, organic molecules with fluorescent properties can be used as in vivo probes. High resolution 3D structures will provide much needed insight into the nature of ligand binding interactions with G-quadruplexes. Currently it is difficult to rationalize ligand affinities due to the absence of appropriate data on geometry of interaction. In order to gain insights into the ligand-quadruplex interaction of individual parts of the structure special attention will be devoted to loop regions. We will also contribute to the development of experimental methods for DNA structure elucidation. Novel ‘in cell’ NMR methods will be developed to facilitate DNA structure elucidation within confines of cell environment.
Significance for the country
Our studies may contribute to development of new areas of research and science in Slovenia, in particular to the studies of G-quadruplex structures that are expected to form in the promoter regions of genes. We expect that our work will allow identification of new structural elements within G-quadruplex structures that may be used as targets to develop new drugs. New small molecule drugs may in turn generate interest within pharmaceutical industry and may ultimately lead to the creation of new jobs in Republic of Slovenia. Novel NMR methodologies that are being developed at the Slovenian NMR centre, which serves a role of regional infrastructural facility, are used in several studies that range from preservation of literature at the national library to identification of objects in museums. NMR centre has been Centre of Excellence since it was awarded this title under FP5 project and has contributed importantly to promotion of Slovenian research and its integration into international exchange and projects. A very important activity envisaged in the proposal is rising awareness of potentials of NMR spectroscopy and in education of younger colleagues at both under- and graduate level.
Most important scientific results
Annual report
2011,
2012,
2013,
final report,
complete report on dLib.si
Most important socioeconomically and culturally relevant results
Annual report
2011,
2012,
2013,
final report,
complete report on dLib.si
