Sequence and pH effects of LNA-containing triple helix-forming oligonucleotides: physical chemistry, biochemistry, and modeling studies.
Triple helix-forming oligonucleotides (TFOs) have been demonstrated to be capable of interfering with gene expression and modifying genomic DNA in a sequence-specific manner. Partial incorporation of 2'-O,4'-C-methylene linked locked nucleic acid (LNA) residues in TFOs has been shown to enhance significantly triple helix formation, whereas the full-length LNA TFO failed to form a stable triplex. This work is aimed at understanding the triple helix-forming properties of LNA-containing TFOs and at optimally designing their sequences. Both DNA thermal melting, gel retardation, and restriction enzyme experiments as well as modeling studies by molecular mechanics were carried out to investigate the base composition/sequence and pH-dependence effects of LNA-containing TFOs, as well as their structural features underlying triple helix formation. Alternating LNA substitution every 2-3 nucleotides in TFOs is mandatory, whereas the use of thymine LNA residues should be favored under neutral pH conditions. A rule for designing optimal LNA-containing TFOs is proposed. In addition, alternative LNA and 2'-O-methyl residues in TFOs do not significantly improve triple helix formation.
Sun
Bei-Wen
BW
Laboratoire de Chimie Structurale et Spectroscopie Biomoléculaire, UMR7033 CNRS, UFR de Santé Medecine Biologie Humaine, Universite Paris-Nord, 93017 Bobigny Cedex, France.
Babu
B Ravindra
BR
Sørensen
Mads D
MD
Zakrzewska
Krystyna
K
Wengel
Jesper
J
Sun
Jian-Sheng
JS
D001483:Base Sequence; D004247:DNA / Q000737:chemistry; D015252:Deoxyribonucleases, Type II Site-Specific / Q000737:chemistry; D006358:Hot Temperature; D006838:Hydrocarbons; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008697:Methane / Q000031:analogs & derivatives* / Q000737:chemistry*; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D009692:Nucleic Acid Heteroduplexes / Q000737:chemistry*; D009838:Oligodeoxyribonucleotides / Q000737:chemistry*; D011685:Purine Nucleotides / Q000737:chemistry; D011742:Pyrimidine Nucleotides / Q000737:chemistry; D015183:Restriction Mapping; D013056:Spectrophotometry, Ultraviolet; D013816:Thermodynamics
D006838:Hydrocarbons; D009692:Nucleic Acid Heteroduplexes; D009838:Oligodeoxyribonucleotides; D011685:Purine Nucleotides; D011742:Pyrimidine Nucleotides; C051802:triplex DNA; C030011:carbene; D004247:DNA; D015252:Deoxyribonucleases, Type II Site-Specific; C035594:TTTAAA -specific type II deoxyribonucleases; D008697:Methane