title: Next-generation sequencing studies guide the design of pyrrole-imidazole polyamides with improved binding specificity by the addition of β-alanine.
abstract: The identification of binding sites for small molecules in genomic DNA is important in various applications. Previously, we demonstrated rapid transcriptional activation by our small molecule SAHA-PIP. However, it was not clear whether the strong biological effects exerted by SAHA-PIP were attributable to its binding specificity. Here, we used high-throughput sequencing (Bind-n-seq) to determine the binding specificity of SAHA-PIPs. Sequence specificity bias was determined for SAHA-PIPs (3 and 4), and this showed enhanced 6 bp sequence-specific binding compared with hairpin PIPs (1 and 2). This finding allowed us to investigate the role of the β-alanine that links SAHA to PIP, and led in turn to the design of ββ-PIPs (5 and 6), which showed enhanced binding specificity. Overall, we demonstrated the importance of β-moieties for the binding specificity of PIPs and the use of cost-effective high-throughput screening of these small molecules for binding to the DNA minor groove.
authors.affiliation: Department of Chemistry, Graduate School of Science, Kyoto University, Kitashirakawa-oiwakecho, Sakyo-ku, Kyoto 606-8502 (Japan).
authors.forename: Chandran
authors.initials: C
authors.lastname: Anandhakumar
authors.forename: Yue
authors.initials: Y
authors.lastname: Li
authors.forename: Seiichiro
authors.initials: S
authors.lastname: Kizaki
authors.forename: Ganesh N
authors.initials: GN
authors.lastname: Pandian
authors.forename: Kaori
authors.initials: K
authors.lastname: Hashiya
authors.forename: Toshikazu
authors.initials: T
authors.lastname: Bando
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Sugiyama
mesh_terms: D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D059014:High-Throughput Nucleotide Sequencing; D007093:Imidazoles / Q000737:chemistry* / Q000378:metabolism; D009757:Nylons / Q000737:chemistry* / Q000378:metabolism; D011758:Pyrroles / Q000737:chemistry* / Q000378:metabolism; D015091:beta-Alanine / Q000737:chemistry* / Q000378:metabolism
chemical_list: D007093:Imidazoles; D009757:Nylons; D011758:Pyrroles; D015091:beta-Alanine; C029899:imidazole; D004247:DNA
keywords: DNA recognition; high throughput screening; next generation sequencing; synthetic biology