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Guiding the Design of Synthetic DNA-Binding Molecules with Massively Parallel Sequencing

2012; American Chemical Society; Volume: 134; Issue: 42 Linguagem: Inglês

10.1021/ja308888c

1943-2984

Jordan L. Meier, Abigail Yu, Ian Korf, David J. Segal, Peter B. Dervan,

Genomics and Chromatin Dynamics

Genomic applications of DNA-binding molecules require an unbiased knowledge of their high affinity sites. We report the high-throughput analysis of pyrrole-imidazole polyamide DNA-binding specificity in a 10(12)-member DNA sequence library using affinity purification coupled with massively parallel sequencing. We find that even within this broad context, the canonical pairing rules are remarkably predictive of polyamide DNA-binding specificity. However, this approach also allows identification of unanticipated high affinity DNA-binding sites in the reverse orientation for polyamides containing β/Im pairs. These insights allow the redesign of hairpin polyamides with different turn units capable of distinguishing 5'-WCGCGW-3' from 5'-WGCGCW-3'. Overall, this study displays the power of high-throughput methods to aid the optimal targeting of sequence-specific minor groove binding molecules, an essential underpinning for biological and nanotechnological applications.