PubMed Health⌕ Search

Biomedical subjects

S I Dimitrov

Publications and source records attributed to S I Dimitrov.

32 records · Page 2Linked to original sources
PubMed1987

Higher order folding of chromatin is induced in different ways by monovalent and by bivalent cations.

The condensation of the 10 nm chromatin filament in the 30 nm fiber by monovalent cations, polyamines and bivalent cations was studied with light scattering at 90 degrees and flow linear dichroism methods. It was found that monovalent cation- and polyamine-induced folding was a two-step process: a precondensation, when a rotation of nucleosomes takes place only, and a condensation step without changes in nucleosome orientation. Divalent cations affected the structure of chromatin in one step only -- condensation of the chromatin filament being accompanied by nucleosome reorientation.

Animals↗
PubMed1987

The globular domain of histone H5 is internally located in the 30 nm chromatin fiber: an immunochemical study.

The location of the globular domain of histone H5 relative to the axis of the 30 nm chromatin fiber was investigated by following the accessibility of this region of the molecule in chicken erythrocyte chromatin to specific antibodies as a function of chromatin structure. Antibodies to the globular domain of H5 as well as their Fab fragments were found to react with chromatin at ionic strengths ranging from 1-80 mM NaCl, the reaction gradually decreasing upon increase of salt concentration. If, however, Fab fragments were conjugated to ferritin, no reaction of the complex with chromatin was observed at salt concentrations higher than 20 mM. The accessibility of the globular part of H5 in unfolded chromatin to the Fab-ferritin complex was also demonstrated with trypsin-digested chromatin. The experiments were carried out by both solid-phase immunoassay and inhibition experiments. The data obtained are consistent with a structure in which the globular domain of H5 is internally located in the 30 nm chromatin fiber.

Animals↗
PubMed1986

Chromatin superstructure. A study with an immobilized trypsin.

Hen erythrocyte chromatin was treated with trypsin immobilized on collagen membranes and the unfolding of chromatin fiber was followed by light scattering at 90 degrees and flow linear dichroism. Chromatin was found almost completely decondensed when the bulk of H1 and H5 was digested while H3 was still intact. Further digestion leading to degradation of both H3 and the rest of H1 and H5 accounted for no more than 10-15% of the total effect. When chromatin with trypsin-cleaved H1 and H5 was titrated with increasing amounts of spermidine it folded similarly to the control sample. This finding suggests that charge neutralization appears a likely mechanism for maintaining the structure of the 30 nm chromatin fiber by the C-terminal domain of H1 and H5.

Animals↗
PubMed1984

The role of histone H1 and non-structured domains of core histones in maintaining the orientation of nucleosomes within the chromatin fiber.

Calf thymus chromatin was digested with trypsin and the structural alterations which occurred were followed by flow linear dichroism. After a sharp initial increase, the amplitude of the positive signal gradually decreased followed by a change of the sign of the dichroism and further increase of the negative signal up to a plateau. These changes of the dichroism were compared to the respective changes in the histone pattern. It was shown that the positive dichroism of chromatin did not depend on the condensation state of chromatin, and that the orientation of the nucleosomes along the chromatin fiber was maintained by the globular domain of H1 and the non-structured parts of core histones.

Animals↗
PubMed1983

Salt-induced conformational transitions in chromatin. A flow linear dichroism study.

The chromatin structure in solution has been studied by the flow linear dichroism method (LD) in a wide range of ionic strengths. It is found that increasing the ionic strength from 0.25 mM Na2EDTA, pH 7.0 to 100 mM NaCl leads to a strong reduction of the LD amplitude of chromatin and inversion of the LD sign from negative to positive at 2 mM NaCl. Chromatin exhibits a positive LD maximum value at 10-20 mM NaCl. These data enable us to conclude that in very low ionic strength (0.25 mM Na2EDTA) the nucleosome discs are oriented with their flat faces more or less parallel to the chromatin filament axis. Increasing ionic strength up to 20 mM NaCl leads to reorientation of the nucleosome discs and to formation of chromatin structures with nucleosome flat faces inclined to the fibril axis. A conformational transition of that kind is not revealed in H1-depleted chromatin. The condensation of the chromatin filaments with increasing concentration of NaCl from 20 mM to 100 mM slightly influences the orientation of the nucleosomes.

Animals↗
PubMed1983

Histone acetylation in chromatin containing mouse satellite DNA.

Histone acetylation in transcriptionally inactive chromatin has been studied with chromatin containing mouse satellite DNA. The latter was obtained by digestion of nuclei from Ehrlich ascites tumor cells with the restriction nuclease Bsp, which degrades main-band DNA but leaves satellite DNA intact. The enzyme-resistant material was separated by gel filtration. Satellite DNA amounted to 65% of the total DNA in this fraction. When the cells were grown in the presence of sodium n-butyrate to inhibit histone deacetylation, a few, if any, hyperacetylated forms of core histones were found in satellite chromatin. Conversely, the highest quantity of tetraacetylated H4 molecules was found in the fractions containing the most extensively degraded chromatin.

Acetylation↗
PubMed1981

Studies on the thermal denaturation of histone-H1-depleted chromatin.

The multiphasic thermal denaturation profile of histone-H1-depleted chromatin was studied by using a nucleoprotein preparation which lacked the first high temperature transition at about 72 degrees C. Such a preparation was obtained by heating at 72 degrees C H1-depleted chromatin, the DNA of which was cross-linked with psoralen to ensure a complete renaturation of DNA upon cooling. When this nucleoprotein was redenatured, its melting profile was found to be significantly altered: only one high temperature peak centered at about 82 degrees C was observed in addition to the low temperature transition at about 53 degrees C. The kinetics of digestion of this material with micrococcal nuclease showed a limit digest equal to that found for the 'native' H1-depleted chromatin but the rate of hydrolysis was higher. The monomer particles prepared from this nucleoprotein were found similar to the 'native' monosomes in respect to protein:DNA ratio and size of DNA but showed an altered melting profile: the premelt area was broader, bigger, and centered at lower temperature; the main peak was reduced in size with no change in its melting temperature. On the basis of these data, the following conclusions were drawn: (a) the last two thermal transitions in H1-depleted chromatin most likely reflect the presence within each nucleosome of two regions with different stability of DNA; (b) DNA involved in the first high thermal transition of H1-depleted chromatin belongs to nuclease-accessible DNA, and (c) the main peak in the biphasic melting profile of the monomer particles reflects the denaturation of DNA regions which in the polymer nucleoprotein are involved in the two high temperature transitions.

Animals↗
PubMed1980

Structural rearrangement of histone-H1-depleted chromatin during thermal denaturation.

The influence of thermal denaturation on the nucleosomal structure of histone-H1-depleted chromatin was studied using psoralen-treated nucleoprotein preparations subjected to partial thermal denaturation. DNA was cross-linked with psoralen to ensure its complete renaturation upon cooling. The structure of the preheated nucleoprotein was investigated by thermal denaturation, kinetics of hydrolysis and DNA fragment pattern obtained upon digestion with micrococcal nuclease. The electron micrographs of the partially denatured nucleohistone showed gross changes in the nucleosomal structure which were consistent with a sliding of histone cores along DNA as recently reported by Tsaneva et al. (Tsaneva, I., Dimitrov, S., Pashev, I. and Tsanev, R., FEBS Lett., (1980) 112, 143-146). This interpretation is strongly supported by the following features of the partially denatured material: a, increased rate of degradation of DNA by micrococcal nuclease; b, melting of a part of DNA as a protein-free DNA; and c, shortening of the DNA repeat length upon digestion with micrococcal nuclease. The sliding of the core histones is parallelled by the denaturation of histones, which accounts for the very intensive background in the DNA digestion pattern, the loss of nucleosome morphology at higher temperatures, and the disappearance in the melting profile of the transition at 72 degrees C.

Animals↗
PubMedDate not supplied

[Study of the process of chromatin condensation using light scattering and stop-flow technics].

We have studied the condensation of calf thymus chromatin induced by NaCl by static light scattering at 90 degrees and showed that the increase of NaCl concentration up to 120-170 mM results in a large increase of scattering intensity of the total chromatin. H1-depleted and trypsinized chromatin preparations do not reveal such a large increase of scattering intensity. The increase of the scattering intensity reflects folding of the chromatin filaments, but not their aggregation. We have used this effect to monitor the kinetics of the chromatin condensation in response to a jump to higher NaCl concentrations by means of a stopped-flow technique. The results show that the condensation is a fast complex process consisting of at least two steps. The first step is only partially resolved by the stopped-flow apparatus. The second step has a time constant in the range of 20-50 ms and does not depend on chromatin concentration.

Animals↗
PubMedDate not supplied

[Linear dichroism of chromatin fibers].

The optical anisotropy of chromatin with different length of the linker DNA isolated from a variety of sources (Friend erythroleukemia cells, calf thymus, hen erythrocytes and sea urchin sperm) has been studied in a large range of mono- and bivalent cations by the use of flow linear dichroism and electric dichroism. We have found that all chromatins studied displayed negative LD values in the range of 0.25 mM EDTA--2 mM NaCl and close positive values in the range of 2-100 mM NaCl. Mg2+ cations, in contrast to Na+ cations, induce optically isotropic chromatin fibers. All chromatin samples exhibit positive form effect amounting to 5-10% of LD amplitude observed at 260 nm. This form effect is determined by the anisotropic scattering of polarized light by single chromatin fibers. The conformational transition at 2 mM NaCl leads to the distortion of chromatin filament structure. The reversibility of this distortion depends on the length of the linker DNA--for chromatins with linker DNA 10-30 b.p. it is partially reversible, while for preparations with longer linker DNA it is irreversible. Relatively low electric fields do not have an effect on chromatin structure, while higher electric fields (more than 7 kV/cm) distort the structure of chromatin.

Animals↗
PubMedDate not supplied

[Interaction of polyamines with chromatin and DNA: formation of compact structures].

We have used flow linear dichroism (LD) and light scattering at 90 degrees to study the condensation of both DNA and calf thymus chromatin induced by spermine, triamines NH3+(CH2)iNH+(CH2)jNH3+, designated as much less than i, j much greater than: much less than 3, 4 much greater than (spermidine), much less than 3, 3 much greater than, much less than 2, 3 much greater than, much less than 2, 2 much greater than; the diamines putrescine and cadaverine and MgCl2. It is found that the different polyamines affected DNA and chromatin in a similar way. The degree of compaction of the chromatin fibers induced by spermine, triamines except much less than 2, 2 much greater than and Mg2+ has been found to be identical. The triamine much less than 2, 2 much greater than and the diamines studied do not condense either chromatin of DNA. Such a big difference in the action of the triamines indicates that not only the charge, but also the structure of the polycations are important for their interactions with DNA and chromatin. The stoichiometry of polyamine binding to chromatin at which condensation occurred is found to be 2 polyamine molecules per DNA helical turn. Polyamines are supposed to bind to the exposed sites of core DNA every 10 b.p. The extent of DNA phosphate neutralization by the histones is estimated to be about 55%. It has been shown that a mixture of mono- and multivalent cations affected DNA and chromatin condensation competitively and not synergistically, as claimed in a recent report by Sen and Crothers.

Animals↗
PubMedDate not supplied

[Study of structural changes in chromatin in the presence of mono- and divalent cations by means of flow linear dichroism].

The chromatin structure in solution was studied by the flow linear dichroism method (LD) in a wide range of ionic strengths. It was found that the increasing of the ionic strength from 0.25 mM EDTA, pH 7.0 to 100 mM NaCl leads to a manifold decrease of the LD amplitude of chromatin and inversion of the LD sign from negative to positive at 2 mM NaCl. Chromatin exhibits a positive LD maximum value at 10-20 mM NaCl. These data enable us to conclude that in very low ionic strength (0.25 mM EDTA-2mM NaCl) the nucleosome discs are oriented with their flat faces parallel to the chromatin filament axis. Increasing ionic strength up to 20 mM NaCl leads to reorientation of the nucleosome discs and to formation of chromatin structure with nucleosome flat faces more or less perpendicular to the fibril axis. A conformational transition of that kind is not revealed in H1-depleted chromatin. This testifies to the fact that the histone H1 is responsible for the higher order structure of chromatin. It was found that divalent cations do not induce the inversion of the LD sign of chromatin.

Animals↗