¹H and ³¹P nuclear magnetic resonance studies of spermine binding to the Z-DNA form of d(m⁵CGm⁵CGm⁵CG)₂. Evidence for decreased spermine mobility

The binding of spermine to the d(m⁵CGm⁵CGm⁵CG) duplex has been studied by proton and phosphorus nuclear magnetic resonance techniques in order to investigate the mobility and nature of spermine bound to the resulting Z-DNA complex. A characterization of the B to Z transition as a function of increasing spermine concentration demonstrated doubling of the non-exchangeable proton and the phosphorus peaks at a ratio of about 1:1 (spermine/duplex) and a re-simplification of the spectrum at 2:1 (spermine/duplex) where about 90% or the DNA was fully converted into the Z-form. However, some of the Z-DNA proton chemical shifts differed between the 1:1 and 2:1 titration points. Since these differences involved primarily the exchangeable terminal imino and amino protons, they could result from end effects. Discrepancies were generally not observed with the non-terminal proton shifts nor with the phosphorus shifts. These proton and phosphorus chemical shift changes are fully consistent with a, B to Z transition. Complexed spermine peaks appear about 0.1 parts per million upfield from the uncomplexed form. The spermine and both the B and Z-DNA hexamer signals are noticeably broadened at the 1:1 ratio but the remaining signals re-sharpen at the 2:1 ratio. Both one-dimensional and two-dimensional studies revealed negative nuclear Overhauser effect (NOE) contacts between each spermine proton. Therefore, spermine has a longer correlation time than that observed for unbounded spermine. These results are contrasted with the positive NOE contacts observed for the B-DNA-spermine complexes reported by Wemmer et al. using the dodecamer d(CGCGAATTCGCG)₂ and reported here using the hexamer d(ATGCAT)₂. While the mobility of spermine in the Z-DNA complex is significantly less than that of the B-DNA complex, no clear evidence of intermolecular spermine-DNA proton NOE contacts is observed.