Overcoming the Challenges in Solution Structure Studies of CTG and CCTG Repeats
研究CTG和CCTG重复序列溶液结构的挑战

Genetic instability of deoxyribonucleic acid (DNA) repeating sequences has been found to associate with hereditary neurodegenerative diseases. It has been suggested that self-expansion of CTG and CCTG repeats would occur through the formation of unusual structures during DNA replication, leading to myotonic dystrophy type 1 (DM1) and myotonic dystrophy type 2 (DM2), respectively. Although x-ray crystallography is a powerful technique in solving DNA structures, there has not been any successful structure determination of CTG or CCTG repeats probably due to the fact that the growth of diffraction-quality crystals has been hampered by the intrinsic flexibility of these repeating sequences. Therefore, nuclear magnetic resonance (NMR) spectroscopy remains to be the only technique that allows studying DNA structures at individual residue level. Recently, we have successfully overcome the challenges in NMR structural studies of CTG and CCTG repeats, including the severe signal overlap due to the repetitive nature of repeating sequences and the increase in spectral complexity owing to the presence of multiple conformers and conformational exchange. With appropriate sample design, sample handling technique, sequence modification and/or single-site substitution experiments, we have determined the structural features of CTG and CCTG repeats using 1H and 31P NMR spectroscopy. This review provides an account of the strategies that have been applied in these studies.