Structural and dynamic differences between supercoiled and linear DNA from proton NMR

Abstract

Proton NMR experiments on an intact plasmid supercoiled DNA molecule are reported. Spectra of the hydrogen-bonded exchangeable imino protons and nonexchangeable aromatic protons of pIns36 (7,200 base pairs; Mr 5 x 10(6)) were examined at 240 and 360 MHz. In contrast to linear DNA of comparable size in which the signals from these protons are broadened almost beyond detection, the supercoiled DNA exhibits a remarkably well-resolved spectrum in the range 12-15 ppm, with linewidths characteristic of linear DNA molecules with molecular weight nearly two orders of magnitude smaller. These results imply an extraordinary degree of mobility in accord with our earlier discovery of unusually long 31P spin-spin relaxation times in the supercoiled molecule. Alterations in the chemical shift of the imino proton resonance positions in supercoiled DNA compared to linear DNA imply that the structure is modified in the A . T base pairs with at least two distinct structures apparent from the spectrum. However, little or no alteration is evident in the resonances due to the G . C base pairs.