Triplex formation of chemically modified homopyrimidine oligonucleotides: thermodynamic and kinetic studies.
We have investigated effects of chemical modifications of a third strand on the thermodynamic and kinetic properties of the triplex formation between a 23-bp duplex and each of four kinds of 15-mer chemically modified third strands using isothermal titration calorimetry and interaction analysis system. The chemical modifications of the third strand included one base modification, with replacement of thymine by uracil; two sugar moiety modifications, RNA and 2'-O-methyl-RNA; and one phosphate backbone modification, with replacement of phosphodiester by phosphorothioate backbone. The thermodynamic and kinetic parameters obtained were similar in magnitude at room temperature for the triplex formation with the base-modified and the sugar-modified third strands. By contrast, binding constant for the triplex formation with the third strand containing phosphorothioate backbone was much smaller by a factor of 10 than that for the other triplex formations. Kinetic analyses have also demonstrated that the third strand containing phosphorothioate backbone was much slower in the association step and much faster in the dissociation step than the other third strands, which resulted in the much smaller binding constant. The reason for the instability of the triplex with the third strand containing phosphorothioate backbone will be discussed. We conclude that, at least in the triplex formation with the chemically modified third strands studied in the present work, the modification of phosphate backbone of the third strand produces more significant effect on the triplex formation than the modifications of base and sugar moiety.
Torigoe
H
H
Gene Bank, Tsukuba Life Science Center, The Institute of Physical and Chemical Research, Tsukuba, Ibaraki 305-0074, Japan. torigoe@rtc.riken.go.jp
Shimizume
R
R
Sarai
A
A
Shindo
H
H
D001483:Base Sequence; D002151:Calorimetry; D066298:In Vitro Techniques; D007700:Kinetics; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000737:chemistry*; D011743:Pyrimidines / Q000737:chemistry; D012965:Sodium Chloride; D013816:Thermodynamics
D009838:Oligodeoxyribonucleotides; D011743:Pyrimidines; D012965:Sodium Chloride