Targeting the weakest links in DNA for selective structural recognition

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Deoxyribonucleic acid (DNA) is unique for each individual, it is linked to genetic disease, and normally forms a double helix. DNA is made up of four bases - adenine (A), thymine (T), guanine (G) and cytosine (C), and with A pairing with T and G pairing with C to give the familiar long chain with a right-handed twist. Nucleic acids (DNA and RNA) can actually adopt a number of different structures, which are greatly dependent on the order of the four bases. The sequence of the bases has a profound effect on the shape and stability of the DNA chain, with specific base steps (e.g. thymine followed by adenine) more flexible than others. Ruthenium polypyridyl complexes are small molecules that have been shown to insert themselves between the bases of DNA and have been investigated for a variety of applications, from light activated anti-cancer therapy to the detection of base mismatches (in which the bases are paired with the incorrect partner and which are linked to the onset of several diseases). These ruthenium complexes are prepared as mixtures of two different molecules (known as enantiomers), that only differ by the direction of the components surrounding the ruthenium centre, like propellers which can turn either way, to the right (clockwise) or to the left (anticlockwise). Much of the research performed on the ruthenium complexes has used an equal mixture of the enantiomers (due to the difficulty in separating the two molecules from one another; they are chemically identical except for this property). Our recent research has shown that the interaction of ruthenium complexes to DNA is not straightforward, with each enantiomer capable of binding to the DNA in a different manner, which could limit their use in the applications mentioned above. We propose to combine our experience in X-ray crystallography (producing solid crystals of DNA and complex that can be used to understand the arrangement of the two components relative to each other) with a systematic study of binding to specific sequences of DNA. We will use enantiomerically pure ruthenium complexes (in which one of the enantiomers is completely separated from the other) to understand how each molecule binds, and the relationship between the two. Experiments conducted in our laboratory, involving crystallography and techniques in solution, have shown that both enantiomers seem to target and bind weakness in the DNA structure and sequence, but sometimes in different ways. This weakness can be in the form of DNA that has been damaged either by the presence of mismatches (where the bases pair with the wrong partner), breaks in the DNA strand, or the presence of bases damaged by environmental conditions (for example chemicals or radiation). Another interesting property of nucleic acids is that they can adopt different structures, distinct from the well known right-twisting double helix. The ability to specifically recognise these alternative structures of nucleic acids will allow us to understand their role within the body, and to develop ways of using them to combat disease. Two of these more specialised structures are the G-quadruplex and i-motif structure, which are four stranded assemblies of DNA that can be formed by specific sequences found in our genetic code. There is a growing body of evidence that suggests that these structures may play important roles in several processes in the body, and being able to target them and selectively stabilise or destabilise could lead to new treatment for diseases as diverse as diabetes to cancer. The proposed research is important in order to unlock the potential of the ruthenium complexes as anti-cancer drugs, or selective biological probes. We must carry out the proposed research to make sense of the way they bind and the effect they then have on the DNA structure and its behaviour in the test tube.