title: Strong, specific, monodentate G-C base pair recognition by N7-inosine derivatives in the pyrimidine.purine-pyrimidine triple-helical binding motif.
abstract: The nucleoside analogs 7-(2'-deoxy-alpha-D-ribofuranosyl)hypoxanthine (alpha7H,1), 7-(2'-deoxy-beta-D-ribofuranosyl)hypoxanthine (beta7H,2) and 7-7-(2'-O-methyl-beta-D- ribofuranosyl)hypoxanthine (beta7HOMe,3) were prepared and incorporated into triplex forming oligodeoxynucleotides, designed to bind to DNA in the parallel (pyrimidine.purine-pyrimidine) motif. By DNase I footprinting techniques and UV-melting curve analysis it was found that, at pH 7. 0, the 15mer oligonucleotides d(TTTTTMeCTXTMeCTMeCTMeCT) (MeC = 5-methyl-deoxycytidine, X =beta7H,beta7HOMe) bind to a DNA target duplex forming a H.G-C base triple with equal to slightly increased (10-fold) stability compared to a control oligodeoxynucleotide in which the hypoxanthine residue is replaced by MeC. Remarkably, triple-helix formation is specific to G-C base pairs and up to 40 microM third strand concentration, no stable triplex exhibiting H.A-T, H.T-A or H.C-G base arrangements could be found (target duplex concentration approximately 0.1 nM). Multiply substituted sequences containing beta7H residues either in an isolated [d(TTTTTbeta7HTbeta7HTbeta7HTbeta7HTbeta7HT)] or in a contiguous [d(TTTbeta7Hbeta7Hbeta7Hbeta7HTTTTbeta7HTTT)] manner still form triplexes with their targets of comparable stability as the control (MeC-containing) sequences at pH 7.0 and high salt or spermine containing buffers. General considerations lead to a structural model in which the recognition of the G-C base pair by hypoxanthine takes place via only one H-bond of the N-H of hypoxanthine to N7 of guanine. This model is supported by a molecular dynamics simulation. A general comparison of the triplex forming properties of oligonucleotides containing beta7H with those containing MeC or N7-2'-deoxyguanosine (N7G) reveals that monodentate recognition in the former case can energetically compete with bidentate recognition in the latter two cases.
authors.affiliation: Department of Chemistry and Biochemistry, University of Bern, Freiestrasse 3, CH-3012 Bern, Switzerland.
authors.forename: J
authors.initials: J
authors.lastname: Marfurt
authors.forename: S P
authors.initials: SP
authors.lastname: Parel
authors.forename: C J
authors.initials: CJ
authors.lastname: Leumann
mesh_terms: D001482:Base Composition; D001483:Base Sequence; D003198:Computer Simulation; D003596:Cytosine / Q000032:analysis*; D004247:DNA / Q000737:chemistry*; D018983:DNA Footprinting; D003850:Deoxyribonuclease I; D004926:Escherichia coli / Q000235:genetics; D006147:Guanine / Q000032:analysis*; D006860:Hydrogen Bonding; D019271:Hypoxanthine / Q000737:chemistry; D007288:Inosine / Q000031:analogs & derivatives* / Q000032:analysis / Q000138:chemical synthesis / Q000737:chemistry; D007364:Intercalating Agents; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000737:chemistry*; D010957:Plasmids; D011687:Purines; D011743:Pyrimidines; D013816:Thermodynamics
chemical_list: D007364:Intercalating Agents; D009838:Oligodeoxyribonucleotides; D011687:Purines; D011743:Pyrimidines; D019271:Hypoxanthine; D007288:Inosine; D006147:Guanine; D003596:Cytosine; D004247:DNA; D003850:Deoxyribonuclease I