Reactive derivatives of oligonucleotide phosphorothioate analogues: IV. Site-directed modification of nucleic acids and intramolecular alkylation of phosphorothioate groups |
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Authors: | N V Amirkhanov N V Neronova V F Zarytova |
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Institution: | (1) Institute of Bioorganic Chemistry, Siberian Division, Russian Academy of Sciences, pr. Akademika Lavrent’eva 8, 630090 Novosibirsk, Russia |
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Abstract: | Alkylation of the 22-mer DNA target pTGCCTGGAGCTGCTTGATGCCC (I) by oligodeoxynucleotide phosphorothioate derivatives (PTAO)
GpsCpsApsTpsCpsApsApsGpsCpsApsGpsCpN(CH3)CH2(RCl)(II-PS) and (RCl)CH2N(CH3)pGpsCpsAps TpsCpsApsApsGpsCpsApsGpsC (III-PS) bearing a residue of an aromatic analogue of nitrogen lost (RCl=C6H4N(CH3)(CH2CH2Cl) at the 3′- or 5′-end was studied. It was shown that the internucleotide phosphorothioate bonds do not affect the regiospecificity
of the target modification. The maximum degree of the target modification (att→∞) at 20°C was about 25% for both (II-PS) and (III-PS). The use of GCATCAAGCAGCpN(CH3)CH2(RCl)(II-PO), containing internucleotide phosphodiester bonds, under the same conditions gave about 65% of the modified DNA.
Kinetics of the PTAO-induced complementarily addressed nucleic acid (NA) modification was analyzed. The rate constants of
the reaction of the intermediate reactive ethylenimmonium ion with phosphorothioate groups of the reagents were evaluated
both in solution and in duplex. The intramolecular alkylation of phosphorothioate groups considerably affected the DNA target
modification by decreasing the effectiveness of the modification in a wide range of temperatures and changing the temperature
dependence of the modification from a bell-like to an S-like profile. It was concluded that, in the course of the modification,
the PTAO phosphorothioate groups are intramolecularly alkylated both in solution and in the complementary NA target-oligonucleotide
duplex.
For Part III, see 1]. |
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Keywords: | alkylating oligonucleotide reagents intramolecular alkylation kinetic equations nucleic acids phosphorothioate analogues site-specific (complementarily addressed) modification |
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