The structural role of histone H1: properties of reconstituted chromatin with various H1 subfractions (H1-1, H1-2, and H1o).

A previous study on the distribution of histone H1 subfractions in chromatin suggested that these proteins differ in the protection they confer to DNA. To elucidate further this suggestion, reconstitution experiments were carried out with purified H1 subfractions (H1-1, H1-2, H1o) and H1-depleted chromatin. We have studied the structural properties of H1o as compared to those of other H1 fractions by electrophoretic analysis of DNA and mononucleosomes obtained after micrococcal nuclease digestion, thermal denaturation, and electron microscopy. The three fractions studied reassociate to H1-depleted chromatin. However, differences in the extent of DNA protection are observed between H1o and the other fractions: H1o induces a more rapid degradation of long oligomers into mononucleosomes; these mononucleosomes bearing H1o only, have a greater electrophoretic mobility; furthermore, thermal denaturation shows that a small fraction of DNA is less efficiently protected by H1o than by the other fractions. Electron microscopy, on the other hand, shows that these differences are not due to areas of chromatin devoid of H1o in the reconstitute and that the reconstituted samples are able, under proper ionic conditions, to refold in a higher-order structure.     

Synthesis of l-(4-2H)erythrose,l-)1-13C, 5-2H)arabinose and l-(2-13C, 5-2H)arabinose and identification of the intermediates by 2H and 13C-N.M.R. spectroscopy

l-(1-¹³C, 5-²H)Arabinose (6D) and l-(2-¹³C, 5-²H)arabinose (8D) have been synthesized by degradation of 2,3-O-isopropylidene-α-l-rhamnofuranose (2) to l-(4-²H)erythrose (5β,5αD), with subsequent chain elongation to 6D plus l-(1-¹³C, 5-²H)ribose (7D), the latter being converted into 8D. Intermediates were identified by complete assignment of the ¹³C chemical shifts employing carbon-carbon and carbon-deuterium coupling constants, deuteration shifts, differential isotope-shifts, and deuterium spectra. The anomeric carbon atoms of 2 and 2,3-O-isopropylidene-l-(1-²H) erythrose (4D) gave only single ¹³C resonances, suggesting that these two compounds exists in only one major anomeric configuration, clarifying previously reported work. The synthesis of 2,3-O-isopropylidene-l-(1-²H)rhmanitol (3D) facilitated the assignment of the signals in the ¹³C spectra of the nondeuterated analog. Specific deuterium-enrichment and the observed carbon-deuterium coupling (¹J_C,D ∼22 Hz) not only served to identify the deuterated carbon atom unambiguously in 3 but also permitted assignment of closely spaced resonances. The deuterium spectrum of 2,3-O-isopropylidene-l-(1-²H)erythrofuranose (4D) showed only a single resonance, indicating preponderance of one anomer, in accord with the observation of a single C-1 resonance in the ¹³C spectrum.