title: Interactions of two large antiviral polyamides with the long control region of HPV16.
abstract: PA1 and PA25 are large hairpin polyamides that are effective in nearly eliminating HPV16 episomes (DNA) in cell culture, and PA25 has broad spectrum activity against three cancer-causing forms of HPV (Edwards, T. G., Koeller, K. J., Slomczynska, U., Fok, K., Helmus, M., Bashkin, J. K., Fisher, C., Antiviral Res. 91 (2011) 177-186). Described here are the interactions of these PAs with sequences in the long control region (LCR) of HPV16 (7348-122). Using an FeEDTA conjugate of PA1 (designed to recognize 5'-W2GW7-3'; W = A or T), 34 affinity cleavage (AC) patterns were detected for this fragment. These sites can be rationalized with sequences featuring perfect, single, double, triple and quadruple mismatches. Quantitative DNase I footprinting analysis indicates that perfect sites bind PA1 with Kds between 0.7 and 2.2 nM. Kds for single, double, triple and quadruple mismatch sites range from 1-3 nM-20 nM. Using AC and EDTA conjugates, we report that unlike smaller 8-ring hairpin PAs, introduction of a chiral turn in this large polyamide has no effect on binding orientation (forward vs. reverse). Despite its design to recognize 5'-W2GW5GW4-3' via two Im residues, a motif not represented in this HPV sequence, a PA25-EDTA conjugate yielded 31 affinity cleavage sites on the region. Low nM Kds for PA25 without EDTA indicates a high tolerance for triple and quadruple mismatches. While there is extensive coverage of the sequence examined, AC cleavage patterns for the two PAs show discrete binding events and do not overlap significantly. This indicates that within the context of A/T rich sequences, these PAs do not recognize a simple shared sequence-related feature of the DNA. These insights continue to inform the complex nature of large hairpin PA-DNA interactions and antiviral behavior.
authors.affiliation: Department of Chemistry & Biochemistry and the Center for Nanoscience, University of Missouri St. Louis, St. Louis, MO 63121, USA.
authors.forename: Elena
authors.initials: E
authors.lastname: Vasilieva
authors.affiliation: Department of Chemistry & Biochemistry and the Center for Nanoscience, University of Missouri St. Louis, St. Louis, MO 63121, USA.
authors.forename: Jacquelyn
authors.initials: J
authors.lastname: Niederschulte
authors.affiliation: Department of Chemistry & Biochemistry and the Center for Nanoscience, University of Missouri St. Louis, St. Louis, MO 63121, USA.
authors.forename: Yang
authors.initials: Y
authors.lastname: Song
authors.affiliation: Department of Chemistry & Biochemistry and the Center for Nanoscience, University of Missouri St. Louis, St. Louis, MO 63121, USA.
authors.forename: George Davis
authors.initials: GD
authors.lastname: Harris
authors.affiliation: Department of Chemistry & Biochemistry and the Center for Nanoscience, University of Missouri St. Louis, St. Louis, MO 63121, USA.
authors.forename: Kevin J
authors.initials: KJ
authors.lastname: Koeller
authors.affiliation: Department of Chemistry & Biochemistry and the Center for Nanoscience, University of Missouri St. Louis, St. Louis, MO 63121, USA.
authors.forename: Puhong
authors.initials: P
authors.lastname: Liao
authors.affiliation: Department of Chemistry & Biochemistry and the Center for Nanoscience, University of Missouri St. Louis, St. Louis, MO 63121, USA.
authors.forename: James K
authors.initials: JK
authors.lastname: Bashkin
authors.affiliation: Department of Chemistry & Biochemistry and the Center for Nanoscience, University of Missouri St. Louis, St. Louis, MO 63121, USA. Electronic address: cdup@umsl.edu.
authors.forename: Cynthia M
authors.initials: CM
authors.lastname: Dupureur
mesh_terms: D000998:Antiviral Agents / Q000378:metabolism*; D001483:Base Sequence; D004279:DNA, Viral / Q000235:genetics* / Q000378:metabolism*; D052162:Human papillomavirus 16 / Q000235:genetics*; D055029:Inverted Repeat Sequences; D009757:Nylons / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics
chemical_list: D000998:Antiviral Agents; D004279:DNA, Viral; D009757:Nylons
keywords: Affinity cleavage; Antiviral; Binding constant; Capillary electrophoresis; DNase I footprinting; Polyamide