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Small-molecule ligand induces nucleotide flipping in (CAG)n trinucleotide repeats

Nature Chemical Biology volume 1, pages 39–43 (2005)Cite this article

Abstract

DNA trinucleotide repeats, particularly CXG, are common within the human genome. However, expansion of trinucleotide repeats is associated with a number of disorders, including Huntington disease, spinobulbar muscular atrophy and spinocerebellar ataxia1,2,3,4. In these cases, the repeat length is known to correlate with decreased age of onset and disease severity5,6. Repeat expansion of (CAG)n, (CTG)n and (CGG)n trinucleotides may be related to the increased stability of alternative DNA hairpin structures consisting of CXG-CXG triads with X-X mismatches7,8,9,10,11. Small-molecule ligands that selectively bound to CAG repeats could provide an important probe for determining repeat length and an important tool for investigating the in vivo repeat extension mechanism. Here we report that napthyridine-azaquinolone (NA, 1) is a ligand for CAG repeats and can be used as a diagnostic tool for determining repeat length. We show by NMR spectroscopy that binding of NA to CAG repeats induces the extrusion of a cytidine nucleotide from the DNA helix.

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Figure 1: Binding of NA to the CAG-CAG triad.
Figure 2: NMR structural analysis of NA-CAG-CAG complex.
Figure 3: NA-CAG-CAG complex.
Figure 4: NA binding to the (CAG)n repeats.
Figure 5: SPR analyses of the (CAG)n repeats by NA-immobilized sensor surface.

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Acknowledgements

We thank T.L. James and M. Shimizu for valuable discussions. This work was partially supported by a Grant in Aid for Scientific Research (A) from the Japan Society for the Promotion of Science to K.N., Health and Labour Sciences Research Grants for Research on Advanced Medical Technology from the Ministry of Health, Labour and Welfare to K.N. and C.K., and CREST, Japan Science and Technology Agency to K.N.

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Author notes

  1. Kazuhiko Nakatani & Masaki Hagihara

    Present address: The Institute of Scientific and Industrial Research (SANKEN), Osaka University, 8-1 Mihogaoka, Ibaraki, 567-0047, Japan

Authors and Affiliations

  1. Department of Synthetic Chemistry and Biological Chemistry, Graduate School of Engineering, Kyoto University, Kyoto, 615-8510, Japan

    Kazuhiko Nakatani, Shinya Hagihara, Yuki Goto & Gosuke Hayashi

  2. PRESTO, Japan Science and Technology Agency, 4-1-8 Honcho Kawaguchi, Saitama, 332-0012, Japan

    Kazuhiko Nakatani, Akio Kobori & Masaki Hagihara

  3. Biotechnology Frontier Project, TOYOBO Co. Ltd., Tsuruga, Fukui, 914-0047, Japan

    Motoki Kyo

  4. Graduate School of Biological Science, Nara Institute of Science and Technology, Nara, 630-0101, Japan

    Makoto Nomura, Masaki Mishima & Chojiro Kojima

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Supplementary information

Supplementary Fig. 1

1H one-dimensional imino proton resonances. (PDF 117 kb)

Supplementary Fig. 2

1H 2D NOESY spectrum of 0.8 mM unlabeled 2:1 NA–CAG–CAG complex. (PDF 170 kb)

Supplementary Fig. 3

NOE constrains used for the structure determination of the 2:1 NA–CAG–CAG complex. (PDF 642 kb)

Supplementary Fig. 4

Discrimination of two possible orientations of NA in the 2:1 complex of NA bound to CAG–CAG triad. (PDF 99 kb)

Supplementary Fig. 5

Confirmation of hairpin formation on (CAG)n upon NA binding. (PDF 168 kb)

Supplementary Table 1

Structural statistics. (PDF 66 kb)

Supplementary Table 2

Detailed 2D and 3D NMR spectral information. (PDF 38 kb)

Supplementary Methods (PDF 78 kb)

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Nakatani, K., Hagihara, S., Goto, Y. et al. Small-molecule ligand induces nucleotide flipping in (CAG)n trinucleotide repeats. Nat Chem Biol 1, 39–43 (2005). https://doi.org/10.1038/nchembio708

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