Fig. 3

Circular dichroism spectra of H1 depleted chromatin and its complexes with linker histone. Polynucleosomes (10–30 mer) were prepared from soluble chromatin of rat testis by ultracentrifugation on a linear 5–40% sucrose density gradient. Linker histone H1 was stripped off using CM Sephadex beads in buffer containing 350 mM NaCl. a Electrophoretic pattern of different fractions of DNA isolated from 5–40% sucrose density gradient ultracentrifugation of MNase digested nuclei in 1.5% agarose gel. Chromatin from 13th–15th fractions (boxed) was used for CD experiments. b 12% SDS-PAGE of proteins associated with native (lane 1) and H1-depleted chromatin (lane 2). c–e, Circular dichroism spectra of histone H1-depleted chromatin after binding with different amount of histone H1 subtype. H1-depleted chromatin was mixed with individual histone subtype in the histone/DNA ratio of 0.0032 (mol/bp); and 0.0064 (mole/bp), respectively, in 80 mM NaCl, 10 mM Tris–HCl, pH 7.4, 0.1 mM EDTA. Spectra were recorded after incubating the chromatin-protein complex overnight at 4 °C. Bar diagram shows the molar ellipticity in different experimental conditions