31P NMR analysis of the DNA conformation induced by protein binding -: SRY/DNA complexes

Complexes of the HMG box protein SRY with two duplexes of 8 and 14 base pairs have been studied by P-31 NMR and complete assignment of all phosphorus signals of the bound DNA duplexes are presented. While for the free DNA, all P-31 signals display Limited spectral dispersion (< 0.8 p.p.m.) for the bound duplexes, P-31 resonances are spread over 2 p.p.m. Based on the previously published 3D structure of hSRY-HMG, with the 8 mer it is demonstrated that the upfield shifted resonances correspond to the site of partial intercalation of an isoleucine side chain into the DNA. Moreover, the observation of significant difference in linewidths between the two duplexes allows to estimate lifetime of the complexes from P-31-P-31 2D exchange experiments.