Recognition of the four Watson–Crick base pairs in the DNA minor groove by synthetic ligands

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Abstract

The design of synthetic ligands that read the information stored in the DNA double helix has been a long-standing goal at the interface of chemistry and biology1,2,3,4,5. Cell-permeable small molecules that target predetermined DNA sequences offer a potential approach for the regulation of gene expression6. Oligodeoxynucleotides that recognize the major groove of double-helical DNA via triple-helix formation bind to a broad range of sequences with high affinity and specificity3,4. Although oligonucleotides and their analogues have been shown to interfere with gene expression7,8, the triple-helix approach is limited to recognition of purines and suffers from poor cellular uptake. The subsequent development of pairing rules for minor-groove binding polyamides containing pyrrole (Py) and imidazole (Im) amino acids offers a second code to control sequence specificity9,10,11. An Im/Py pair distinguishes G·C from C·G and both of these from A·T/T·A base pairs9,10,11. A Py/Py pair specifies A,T from G,C but does not distinguish A·T from T·A9,10,11,12,13,14. To break this degeneracy, we have added a new aromatic amino acid, 3-hydroxypyrrole (Hp), to the repertoire to test for pairings that discriminate A·T from T·A. We find that replacement of a single hydrogen atom with a hydroxy group in a Hp/Py pairing regulates affinity and specificity by an order of magnitude. By incorporation of this third amino acid, hydroxypyrrole–imidazole–pyrrole polyamides form four ring-pairings (Im/Py, Py/Im, Hp/Py and Py/Hp) which distinguish all four Watson–Crick base pairs in the minor groove of DNA.

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Figure 1: Chemical structures and space-filling models of the T·A and A·T base pairs as viewed from the minor groove of DNA.
Figure 2: Eight-ring hairpin polyamides containing three aromatic amino acids (Py, Hp, Im).
Figure 3: Quantitative DNase I footprint titration experiments.

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Acknowledgements

We are grateful to the NIH for research support and National Research service Awards to S.W. and J.W.S., to the NSF for a predoctorial fellowship to S.W., to J. Edward Richter for an undergraduate fellowship to J.M.T., and to the HHMI for a predictorial fellowship to E.E.B.

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Correspondence to Peter B. Dervan.

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