Positively charged deoxynucleic methylthioureas: Synthesis and binding properties of pentameric thymidyl methylthiourea

The negatively charged phosphodiester linkages in DNA {-O(PO2-)O-} have bee n replaced by a novel methylthiourea {-NHC(=SMe+)NH-} backbone. The backbon e is positively charged, achiral, and stable and can be easily synthesized. A basic strategy for the synthesis of deoxynucleic methylthioureas (DNmt) is described. Synthetic procedures are provided for thymidyl DNmts (1-4 lin kages). Synthesis proceeds in 3'-5' direction and involves coupling of a pr otected 3'-isothiocyanate with the corresponding S-amine of the growing oli go chain. Coupling reactions at room temperature are nearly quantitative, a nd Products are easily purified. The method of continuous variation indicat es that there is an equilibrium complex with a molar ratio of d(Tmt) to r(A p) or d(Ap) of 2:1. Continuous variation plots carried out at temperatures from 15 to 60 degrees C show a triple helical complex. Titration scans over the entire range of wavelengths (240-285 nm) confirm binding in triple hel ical fashion. Thermal denaturation analyses show pronounced hysteresis with poly(dA) as well as poly(rA). Hysteresis is more pronounced at higher ioni c strengths due to a slower annealing process. DNmt shows fidelity in bindi ng to polynucleotides as there is little hyperchromicity observed in denatu ration of DNmt complexes to noncomplementary deoxynucleotides and ribonucle otides. The effect of ionic strength on thermal denaturation is very pronou nced, with stability greatest at low ionic strengths. Thermal denaturation studies show melting points of >15 degrees C per base pair in complexes of DNmt with poly(da) and poly(rA).