Poly(L-lysine)-graft-dextran copolymer promotes pyrimidine motif triplex DNA formation at physiological pH. Thermodynamic and kinetic studies.
Extreme instability of pyrimidine motif triplex DNA at physiological pH severely limits its use for artificial control of gene expression in vivo. Stabilization of the pyrimidine motif triplex at physiological pH is therefore of great importance in improving its therapeutic potential. To this end, isothermal titration calorimetry interaction analysis system and electrophoretic mobility shift assay have been used to explore the thermodynamic and kinetic effects of our previously reported triplex stabilizer, poly (L-lysine)-graft-dextran (PLL-g-Dex) copolymer, on pyrimidine motif triplex formation at physiological pH. Both the thermodynamic and kinetic analyses have clearly indicated that in the presence of the PLL-g-Dex copolymer, the binding constant of the pyrimidine motif triplex formation at physiological pH was about 100 times higher than that observed without any triplex stabilizer. Of importance, the triplex-promoting efficiency of the copolymer was more than 20 times higher than that of physiological concentrations of spermine, a putative intracellular triplex stabilizer. Kinetic data have also demonstrated that the observed copolymer-mediated promotion of the triplex formation at physiological pH resulted from the considerable increase in the association rate constant rather than the decrease in the dissociation rate constant. Our results certainly support the idea that the PLL-g-Dex copolymer could be a key material and may eventually lead to progress in therapeutic applications of the antigene strategy in vivo.
Gene Bank, Tsukuba Life Science Center, The Institute of Physical and Chemical Research (RIKEN), 3-1-1 Koyadai, Tsukuba, Ibaraki 305-0074, Japan. torigoe@rtc.riken.go.jp
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H
Torigoe
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Ferdous
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H
Watanabe
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Akaike
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Maruyama
D004247:DNA; D003911:Dextrans / Q000494:pharmacology*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009690:Nucleic Acid Conformation / Q000187:drug effects*; D011107:Polylysine / Q000031:analogs & derivatives* / Q000494:pharmacology; D011743:Pyrimidines; D013816:Thermodynamics
D003911:Dextrans; D011743:Pyrimidines; C113090:poly(lysine)-graft-dextran; C051802:triplex DNA; D011107:Polylysine; D004247:DNA; C030986:pyrimidine