DNA supercoiling by core histone fractions and protamine.

The nuclear extract isolated from late Drosophila melanogaster embryos has Mg2+-dependent DNA topoisomerase 1 and nuclease activities. The extract facilitates the closed circular duplex DNA supercoiling in the presence of calf histone fractions H2A, H2B, H3 and H4, and fish protamine but not HI histone.

[Unfolding of nucleosome cores induced by chemical acetylation of histones]. / Razvorachivanie korovykh nukleosom, vyzvannoe khimicheskim atsetilirovaniem gistonov.

Relative accessibility of nucleosomal histones to acetic anhydride during acetylation has been studied as a function of concentration, pH and ionic strength of the solution using high-resolution gel-electrophoresis. It was shown that about 80% of lysine residues in nucleosomal histones and 100% of the same residues in histone complexes without DNA in 2 M NaCl are accessible to the modification, which is proved by the localization of the majority of lysine residues in nucleosomes near the surface of the histone octamer, by their participation in ionic interactions with DNA and, probably, in histone-histone contacts. Gel-electrophoretic experiments with nucleosomes and studies of the histone resistance to mild trypsinolysis indicated that neither nucleosomes themselves nor histone octamers are affected even though 50% of lysine residues in histones have been acetylated. The process of acetylation is accompanied by the growing tendency of histones to participate in mild trypsinolysis and by a gradual decline in electrophoretic mobility and in the value of the sedimentation constant. The circular dichroism spectra and the microscopic appearance of nucleosomes are also markedly changed. These results suggest that a gradual unfolding of nucleosomes occurs when 5 or more lysine residues in the nucleosomal histones have been acetylated.

[Chemical acetylation, trypsinolysis and stability of nucleosomes]. / Khimicheskoe atsetilirovanie, tripsinoliz i stabil'nost' nukleosom.

Gel electrophoretic analysis of the histone chemical acetylation in the nucleosome core particles with acetic andydride revealed availability of about 14 lysine residues of histone H2A, 15-21 of H2B, 8-11--H3 and 6-9--H4. Moderately lysine-rich histones H2A and H2B were found to be more susceptible to acetylation than arginine-rich H3 and H4. Chemical acetylation enhanced the rate of trypsin digestion in acetylated nucleosomes as evidenced by gel electrophoresis of histone fragments. A more pronounced trypsin digestion was evident at acetylation of only 3-5 histone amino groups per nucleosome. However, even heavily acetylated nucleosomes yielded in familiar trypsin limit digest pattern of histone fragments thus indicating persistence of histone octamer. Nucleosomes which were trace acetylated (up to 3-5 histone amino groups neutralized per nucleosome) and treated with trypsin to remove highly charged terminal histone regions revealed remarkable unfolding and partial dissociation when analyzed by gel electrophoresis. The same trace acetylated nucleosomes did not show such destabilization prior to trypsin digestion.

[Complexes of histone H1 with supercoiled DNA]. / Issledovanie kompleksov gistona H1 s superspiral'noi DNK.

The action of DNA topoisomerase I on the complexes of supercoiled DNA (DNA I) with histone H1 and other histones was studied at different ionic conditions and histone/DNA ratios. In the presence of 0.15 M NaCl at histone H1/DNA ratios lower than 0.25 (w/w) the relaxation of DNA I is not inhibited. Raising of H1/DNA I ratio up to 0.7 is followed by significant inhibition of DNA I relaxation. At fixed H1/DNA I ratio maximal inhibition is detected at 0.25 M NaCl. At NaCl concentrations lower than 0.1 M and higher than 0.3 M increasing of DNA I relaxation in the presence of H1 was observed. Electron microscopic studies show that increase of ionic strength or H1/DNA I ratio causes more dense packing of DNA molecules in the H1.DNA complexes. Changes in the structure of complexes agree with the increase of DNA I relaxation inhibition in these conditions. DNA I relaxation inhibition by H1 is drastically decreased by iodination of tyrosine 72 residue in the globular part of H1 molecule. Individual core histones inhibit DNA I relaxation at much higher histone/DNA ratios and show different dependence of inhibition on ionic strength. The results are discussed in terms of the possible role of H1 in chromatin condensation-decondensation.

[Drosophila melanogaster embryo factor capable of supercoiling circular covalently closed DNA in the presence of core histones H2a, H2b, H3, H4 or protamine]. / Superspiralizatsiia kol'tsevoi kovalentno zamknytoi DNK v prisutstvii gistonov H2, H2b, H3, H4 ili protamina i ekstrakta iz embrionov Drosophila melanogaster.

The Mg2+-dependent factor capable of introducing the superhelical turns into circular closed DNA in the presence of core histones H2a, H2b, H3, H4 or protamine in solution at physiological conditions has been isolated from Drosophila melanogaster embryos. The factor does not work with histone H1, cytochrome c, poly-L-lysine or tetralysine. It is assumed that the factor is responsible for folding of core histones into nucleosomes.