The DNA molecule is a carrier of genetic information and a subject of extensive research. Besides the well-known Watson-Crick structure, there exist different DNA structures, for example G-quadruplexes and i-motifs. They appear in important regions in the genome and participate in processes such as replication and transcription. They are often found in the promoter regions of oncogenes and are extensively studied as possible drug targets in cancer treatment.
In this thesis we focused on folding of a short DNA sequence into i-motif. For its formation, the protonation of cytosine and the emergence of C+-C pair is necessary. If the sequence is cytosine-rich, several such pairs may be formed and the result is folding of the chain into i-motif.
The subject of this research is folding and unfolding of cytosine-rich sequence DNA
(5'-CCTTCCCCACCCTCCCCACCCTCA-3') to i-motif. The sequence is found in the promoter of human proto-oncogene c-myc, whose increased expression is associated with a large number of cancers. With a combination of spectroscopic and calorimetric measurements we investigated the thermodynamics of folding and unfolding, which was achieved by regulating pH and temperature. It turned out that folding, achieved by modifying pH is reversible process, while cooling the heat-denatured i-motif doesn’t lead to the initial i-motif structure. We found that binding of protons is not completely cooperative. For the observed change of conformation, it appears, that only two or three of the five expected cytosine pairs are needed for folding. The enthalpy of folding to
i-motif was estimated by taking into account two contributions, namely the establishment of π-π interaction between stacked bases of unfolded chain and further folding to i-motif. Process is governed by negative change in enthalpy. Entropy and heat capacity change accompanying folding are also negative.