High-field NMR and restrained molecular modeling studies on a DNA heteroduplex containing a modified apurinic abasic site in the form of covalently linked 9-aminoellipticine.
Two-dimensional NMR methods were used to model the possible solution structure of an intercalative complex of 9-aminoellipticine (Aell), a polycyclic pyridocarbazolamine, covalently bound to an apurinic ring-opened deoxyribose site of a duplex DNA fragment in the reduced Schiff base form. The required oligonucleotide single strand containing covalently attached aminoellipticine was obtained by reductive amination in the presence of sodium cyanoborohydride. The combined NMR-energy minimization methods were employed to refine the model structures of two distinct forms, intrahelical and extrahelical, of a control 9-mer duplex DNA, d(CGTG.dr.GTGC).d(GCACTCACG), which contains an apurinic site positioned opposite a dT residue on the complementary strand. The model structure of an aminoellipticine conjugate with the same DNA sequence, derivatized via the aforementioned covalent attachment, was also obtained by incorporating intermolecular drug-DNA and intra- and internucleotide NOE-derived proton-proton distance estimates as restraints in energy minimization routines. The indole ring system of aminoellipticine, which is inserted at the apurinic site, intercalates between and is parallel to flanking GC base pairs. The pyridinic ring of aminoellipticine, in protonated form, also stacks between cytidine and thymidine bases on the complementary strand, which is consistent with the observation that the normal sequential NOE connectivity at the 5'-C13-T14 step is broken and indeed diverted through the ellipticine moiety, e.g., C13-Aell-T14 connectivities through the Aell-H4/C5Me protons. Interestingly, the partial stacking of the pyridinic ring is observed only between the 5'-CT step vs an adjacent 5'-TC step, owing to inherently weak stacking interactions associated with the former. In the absence of any potential groups that can participate in electrostatic or hydrogen-bonding interactions with the nucleic acid, pi-pi stacking and hydrophobic contacts at the intercalation site appear to be the important factors in determining stability and conformation of the aminoellipticine-DNA conjugate. Stacking interactions in such a bistranded intercalative complexation of aminoellipticine apparently govern the formation of a single intrahelical form of a right-handed B-type DNA duplex. The overall structural features lead us to propose working models for an enzyme-like DNA cleavage activity of 9-aminoellipticine and the observed inhibition of the AP endonuclease-dependent DNA excision-repair pathway.
Singh
M P
MP
Department of Chemistry, University of Alberta, Edmonton, Canada.
Hill
G C
GC
Péoc'h
D
D
Rayner
B
B
Imbach
J L
JL
Lown
J W
JW
D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000138:chemical synthesis / Q000737:chemistry*; D004260:DNA Repair / Q000187:drug effects; D004611:Ellipticines / Q000737:chemistry* / Q000494:pharmacology; D007364:Intercalating Agents / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009692:Nucleic Acid Heteroduplexes / Q000138:chemical synthesis / Q000737:chemistry*; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000737:chemistry; D012545:Schiff Bases / Q000737:chemistry
D004611:Ellipticines; D007364:Intercalating Agents; D009692:Nucleic Acid Heteroduplexes; D009838:Oligodeoxyribonucleotides; D012545:Schiff Bases; C038373:9-aminoellipticine; D004247:DNA