Infrared spectroscopy of nucleoprotein complexes

Fourier transform infrared (FT-IR) spectroscopy was used in the study of a secondary structure of the DNA-bound Histone H1 (CTD) and also of protamine inside sperm nuclei. The carboxy-terminal domain of histone H1 was measured at 5 mg mL 1 and the appropriate amounts of DNA in 10 mM HEPES buffering agent, pH 7, plus 10 mM or 140 mM NaCl. Salmon and squid sperm nuclei samples were measured at equivalent DNA concentrations of 5 mg mL 1 and 25 mg mL 1, in 10 mM HEPES plus 140 mM NaCl, pH 7. Samples were exchanged with D2O using successive steps of incubation with deuterated buffer. The number and position of the amide I band components were obtained by Fourier deconvolution and used for the curve fitting of the original envelope by an iterative process. The spectra of the complexes of the CTD with DNA were recorded both in H2O and D2O to facilitate the assignment of the amide I components.