Triplex Formation at Physiological pH by 5-Me-dC-N4-(Spermine) [X] Oligodeoxynucleotides: Non Protonation of N3 in X of X*G:C Triad and Effect of Base Mismatch/Ionic Strength on Triplex Stabilities

Abstract

Oligodeoxynucleotide (ODN) directed triplex formation has therapeutic importance and depends on Hoogsteen hydrogen bonds between a duplex DNA and a third DNA strand. T*A:T triplets are formed at neutral pH and C+*G:C are favoured at acidic pH. It is demonstrated that spermine conjugation at N4 of 5-Me-dC in ODNs 1-5 (sp-ODNs) imparts zwitterionic character, thus reducing the net negative charge of ODNs 1-5. sp-ODNs form triplexes with complementary 24mer duplex 8:9 show foremost stability at neutral pH 7.3 and decrease in stability towards lower pH, unlike the normal ODNs where optimal stability is found at an acidic pH 5.5. At pH 7.3, control ODNs 6 and 7 carrying dC or 5-Me-dC, respectively, do not show any triple helix formation. The stability order of triplex containing 5-Me-dC-N4-(spermine) with normal and mismatched duplex was found to be X*G:C approximately X*A:T > X*C:G > X*T:A. The hysteresis curve of sp-ODN triplex 3*8:9 indicated a better association with complementary duplex 8:9 as compared to unmodified ODN 6 in triplex 6*8:9. pH-dependent UV difference spectra suggest that N3 protonation is not a requirement for triplex formation by sp-ODN and interstrand interaction of conjugated spermine more than compensates for loss in stability due to absence of a single Hoogsteen hydrogen bond. These results may have importance in designing oligonucleotides for antigene applications.

(2S,5R/2R,5S)-Aminoethylpipecolyl aepip-aegPNA chimera: synthesis and duplex/triplex stability

“…The oligomers after solid phase assembly were cleaved from the support by treatment with TFA-TFMSA 16 to yield the corresponding PNAs carrying b-alanine at the carboxy terminus. PNAs (13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26) were purified by FPLC on a PepRPC column, and the purity of the oligomers was rechecked by HPLC on RPC-18 column and these were characterized by MALDI-TOF mass spectrometry. 17 The modified aeg-aepipPNA oligomers 13, 14, 18-21 permit the study of the positional effects on PNA 2 :DNA triplex stability, while the mixed base PNA sequences 25-26 allow explicit testing of the relative stereochemical effects of (2S,5R) and (2R,5S) aepipPNA units on duplex formation.…”

“…The compounds were visualized with UV light and/or by spraying with Ninhydrin reagent subsequent to Boc-deprotection (exposing to HCl vapors) and heating. The DNA oligomers were synthesized on CPG solid support by b-cyanoethyl phosphoramidite chemistry followed by ammonia treatment 21 and their purities checked by HPLC prior to use. aegPNA monomers were synthesized according to literature procedures.…”

Dipyrido[3,2-a:2′,3′-c]phenazine-Tethered Oligo-DNA: Synthesis and Thermal Stability of Their DNA⋅DNA and DNA⋅RNA Duplexes and DNA⋅DNA⋅DNA Triplexes

“…Thermal Denaturation Studies of Triplexes. Triplexes (Entries 2 ± 13 in Table 3) were generated by hybridization of the dppz-tethered 18-mers 48 ± 56 and dppztethered 17-mers 57 ± 59 with the target 24-mer duplex D24´D24 (62) [28] (18-mer (or 17-mer)/D24/D24 1 : 1 : 1; 1 mm of each strand in 20 mm PO 4 3À and 0.1m NaCl buffer at pH 7.3). Three similar transitions were observed for all melting profiles.…”

Impact of the Guanidinium Group on Hybridization and Cellular Uptake of Cationic Oligonucleotides

“…Along the same lines, it was anticipated that ON analogues carrying a net positive charge should even be more beneficial in terms of binding efficiency and should also give hybridization with increased association rates. A few examples of cationic ONs (with positively charged backbones) [8][9][10][11][12] or zwitterionic ONs (with positively charged tethers attached to the nucleobases or to the 2'-position of the sugars) [13][14][15][16][17][18][19][20][21][22] have been described. Strategies have included the incorporation of amino groups prone to protonation under physiological conditions or of the guanidinium group (pK a 12.5), which is highly basic and positively charged over a wide pH range.…”