NMR studies on the B-Z transition of synthetic DNA d[(CG)3TA(CG)3]

Abstract

The formation of Z-DNA has been demonstrated in some sequences which are alternating purines and pyrimidines and which contains A·T as well as G·C base pairs. But the $d(A-T)_n$ stretch disrupts the formation of Z-DNA, we have studied about the conformation of DNA oligomer, $d[(CG)_3TA(CG)_3]$ (TA 14mer) which composes of two d(CG)3 stretches and the middle d(TpA) step, using one- and two- dimensional $^1H$ NMR, $^31P$ NMR and CD spectroscopy. We can see that the TA 14mer forms the B-form DNA duplex in 0.2 M NaCl, 10 mM phosphate buffer, pH 7.0, aqueous solution using the $^1H$, $^31P$ NMR and CD spectroscopy. The chemical shifts of sugar H1``, H2``/H2", and base proton resonances for B-form TA 14mer are assigned using NOESY spectrum. While the CD and $^1H$ NMR spectra of TA 14mer in 2 mM $Co(NH_3)_6Cl_3$ and 20 mM NaCl show that the conformation of TA 14mer is B-form at low temperature (20-30℃) but Z-form at high temperature (40-50℃), and that proportion of Z-form depends on the temperature. The resonances of the GpC phosphate shift to downfield in the $^31P$ NMR spectrum at 50℃ and the strong crosspeaks between GH1`` and GH8 are observed in NOESY spectrum at 30℃. From these results, we can confirm that the conformation of the TA 14mer changes to Z-form at high temperature. The chemical shifts of the base proton resonances for the Z-form TA 14mer are assigned by the exchange NOESY spectrum at 50℃ and compared with that of B-form. Comparison of integrated intensities of two $^31P$ resonances for Z-form TA 14mer gives us that the middle TpA step is in the Z-conformation and overall structure of this oligomer is the Z-Z junction. In summary, the polyvalent cation, $Co(NH_3)_6^{3+}$, in millimolar concentration induces the B-Z transition at high temperature and TA 14mer forms the Z-Z junction under this condition.