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J Ausio

Publications and source records attributed to J Ausio.

46 records · Page 3Linked to original sources
PubMed1987

Transition of chromatin from the "10 nm" lower order structure, to the "30 nm" higher order structure as followed by small angle X-ray scattering.

Chromatin oligomers undergo a conformational change from a "10 nm" lower order structure at low concentration of salt to a "30 nm" higher order structure, with increasing NaCl or MgCl2 concentration. We have extended our previously reported hydrodynamic and light-scattering measurements of the folding of well-defined chicken erythrocyte chromatin fractions to include a study of the low angle X-ray scattering in solution. We show that it is feasible to identify the folding process with gradual compaction of a chain of freely joined filaments or a worm-like chain, within the limits of all the experimental data obtained. As the ionic strength is raised, the filament length of the oligomer, composed of Nz nucleosomes, decreases. At 75 mM-NaCl, the compacted model chains (Nz = 53) form structures that are, on average, cylindrically shaped with mean diameter 30 nm and length 104 nm. Helical symmetry need not be invoked in the modelling of the folding process and may, in particular, be difficult to establish in chicken erythrocyte chromatin, due to the non-uniform length of the DNA linker connecting the nucleosomes. Concerning the shape of the X-ray scattering profiles at various salt concentrations, it is possible in this way to rationalize two-slope cross-sectional plots, which have also been reported by other workers. Though this description represents a satisfactory conceptual presentation of a wealth of experimental data, it by no means represents a definitive solution to an exceedingly difficult problem.

Animals↗
PubMed1986

Biochemical and physiochemical characterization of chromatin fractions with different degrees of solubility isolated from chicken erythrocyte nuclei.

Chicken erythrocyte chromatin was prepared according to two different methods [Fulmer, A. W., & Bloomfield, V. A. (1981) Proc. Natl. Acad. Sci. U.S.A. 78, 5968-5972; Ausio, J., Borochov, N., Seger, D., & Eisenberg, H. (1984) J. Mol. Biol. 177, 373-398] to give three main common fractions, according to its solubility (S) or insolubility (I) in 0.15 M NaCl buffers or to its further solubility in 0.25 mM ethylenediaminetetraacetic acid (E). From the biochemical point of view, all of them have been found to be undistinguishable. Analytical ultracentrifugation shows that all of these fractions can reversibly undergo the transition from the low to the higher order structure, through a nearly identical way of folding. Thermal denaturation profiles yielded three transitions having the same Tm's for the three fractions. The percentage of DNA melting in the first transition decreased in the order S greater than I greater than E, and the amount in the second transition increased in the same order. Together with the different solubility of these fractions in the presence of divalent ions, these results indicate that in the three fractions of chromatin studied, the amount of linker DNA bound to the nucleosome varied.

Animals↗
PubMed1986

Histone hyperacetylation: its effects on nucleosome conformation and stability.

We have prepared nucleosome particles from HeLa cells that have been subjected to butyrate treatment. Fractions containing different levels of acetylation have been obtained within the range 7-17 acetyl groups per nucleosome. We have put special emphasis in the characterization of the particles with the highest level of histone acetylation. At low to physiological ionic strengths, these nucleosomes exhibit only small differences in hydrodynamic behavior and circular dichroism from control particles with minimal acetylation. There are, however, significant differences in thermal denaturation and nuclease sensitivity. In terms of stability toward high salt, the hyperacetylated and control particles behave identically. A model that reconciles these results is proposed. The major conclusion from our results, however, is that, at physiological ionic strength and in the absence of factors other than acetylation, the highly hyperacetylated nucleosomes remain essentially folded.

Acetylation↗
PubMed1985

Nucleosome core particle structure and structural changes in solution.

The radius of gyration, Rg, of chicken erythrocyte nucleosome core particles, was found to be 4.56 (+/- 0.07) nm by small-angle X-ray scattering, independent of particle concentration and of NaCl concentration between 0.1 M and 0.6 M-NaCl. The large, positive, second virial coefficient, A2, from particle concentration dependence (but independent of NaCl concentration) in small-angle X-ray scattering may indicate non-electrostatic repulsive ordering over large distances. Density contrast variation in equilibrium sedimentation with a small probe (sucrose) and a larger probe (gamma-cyclodextrin) yields good results for core particle hydration in the first instance, and for an estimate of the total particle volume in the second instance.

Animals↗
PubMed1984

Interaction of chromatin with NaCl and MgCl2. Solubility and binding studies, transition to and characterization of the higher-order structure.

Chicken erythrocyte chromatin containing histones H1 and H5 was carefully separated into a number of well-characterized fractions. A distinction could be made between chromatin insoluble in NaCl above about 80 mM, and chromatin soluble at all NaCl concentrations. Both chromatin forms were indistinguishable electrophoretically and both underwent the transition from the low salt "10 nm" coil to the "30 nm" higher-order structure solenoid by either raising the MgCl2 concentration to about 0.3 mM or the NaCl concentration to about 75 mM. The transitions were examined in detail by elastic light-scattering procedures. It could be shown that the 10 nm form is a flexible coil. For the 30 nm solenoid, the assumption of a rigid cylindrical structure was in good agreement with 5.7 nucleosomes per helical turn. However, disagreement of calculated frictional parameters with values derived from quasielastic light-scattering and sedimentation introduced the possibility that the higher-order structure, under these conditions, is more extended, flexible, or perhaps a mixture of structures. Values for density and refractive index increments of chromatin are also given. To understand the interaction of chromatin with NaCl and with MgCl2, a number of experiments were undertaken to study solubility, precipitation, conformational transitions and binding of ions over a wide range of experimental conditions, including chromatin concentration.

Animals↗
PubMed1984

Nucleosome core particle stability and conformational change. Effect of temperature, particle and NaCl concentrations, and crosslinking of histone H3 sulfhydryl groups.

We have studied the reversible dissociation of core size DNA from chicken erythrocyte nucleosome core particles in solutions containing 0 X 1 M to 0 X 6 M-NaCl. Dissociation increases with increasing NaCl concentration, increasing temperature and decreasing particle concentration. At high particle concentrations, no free DNA is observed below 0 X 3 M-NaCl, whereas above 0 X 3 M-NaCl a lower limit of dissociation is reached. A theoretical analysis based on the migrating-octamer mechanism of Stein is in disagreement with his conclusions concerning dependence of core particle dissociation on particle concentration, but provides a good explanation for our observations, and those of others, using salt concentrations up to 1 M-NaCl. It appears that the core particle is not stabilized primarily by electrostatic interactions. DNA length is not critical for core particle stabilization. The conformation of remaining intact nucleosome core particles changes only moderately within the range of NaCl concentrations studied. Crosslinking by copper phenanthroline of the Cys110 histone H3 single sulfhydryl groups in the intact nucleosome core particle leads to a decrease in stability, yet essentially unchanged hydrodynamic properties are maintained at 0 X 6 M-NaCl, confirming conclusions derived from the behavior of the native core particles. Values for density increments of nucleosome core particles over a range of NaCl concentrations are also given. A method is described for studying binding of histones to nucleosome core particles in the ultracentrifuge by scanning at 230 and 260 nm.

Centrifugation, Density Gradient↗
PubMed1984

Interaction and conformational changes of chromatin with divalent ions.

We have investigated the interaction of divalent ions with chromatin towards a closer understanding of the role of metal ions in the cell nucleus. The first row transition metal ion chlorides MnCl2, CoCl2, NiCl2 and CuCl2 lead to precipitation of chicken erythrocyte chromatin at a significantly lower concentration than the alkali earth metal chlorides MgCl2, CaCl2 and BaCl2. A similar distinction can be made for the compaction of chromatin to the "30 nm" solenoid higher order structure which occurs at lower MeCl2 concentration in the first group but at the same MeCl2 concentration within each group. In other experiments in which mixed solutions of NaCl and of MgCl2 were examined, it is shown that increasing NaCl concentration leads to increasing solubility in the presence of MgCl2. Best compaction of chromatin was obtained at 40 mM NaCl and 0.8 mM MgCl2 at a value A260 approximately 0.8. Similar experiments were undertaken with mixtures of NaCl and MnCl2.

Animals↗