Physical properties of inner histone-DNA complexes

Abstract

Chicken-erythrocyte inner histone tetramer has been complexed with several natural and synthetic DNA duplexes by salt-gradient dialysis at various protein/DNA ratios. The resulting complexes, in low-ionic-strength buffer, have been examined by electron microscopy, circular dichroism, and thermal denaturation. Electron microscopy reveals nucleosomes (v bodies) randomly arranged along DNA fibers, including poly(dA-dT)*poly(dA-dT), poly(dI-dC)*poly(dI-dC), but not poly(dA)*poly(dT).Circular dichroism studies showed prominent histone or-helix and "suppression" of nucleic acid ellipticity (X>240 nm). Thermal denaturation experiments revealed Tm behavior comparable to that of HI-(or H5-) depleted chromatin. Tm III and Tm IV increased linearly with G+C% (natural DNAs), but were virtually independent of the histone/DNA ratio; therefore, the melting of nucleosomes along a DNA chain is insensitive to adjacent "spacer" DNA lengths. This suggests that Tm III and Tm IV arise from the melting of different domains of DNA associated with the core v body.

Conformation of DNA in chromatin reconstituted from poly [d(A-T)] and the core histones

“…This region appears to be devoid of nucleosomes as shown by electron microscopy (8)(9)(10). In vitro experiments have shown that using purified histones H2A, H2B, H3 and H4, the reconstitution of nucleosomes is possible with polydeoxynucleotides composed of alternating purine- (11,12,13). We are thus prompted to ask the question as to the structural features which allow particular DNA sequences to reconstitute nucleosomes.…”

Studies on the Thermal Denaturation of Histone-H1-Depleted Chromatin

“…The following possibilities were suggested [ 5 ] : (a) the presence within each nucleosome of two regions with different thermal stability of DNA, (b) the existence of two structurally different classes of nucleosomes, and (c) the melting of nucleosomal DNA depends on the length of the adjacent spacer regions. The last possibility was ruled out by Bryan et al [12] in experiments with reconstituted histone-DNA complexes. They showed also that the existence of the two transitions could not be explained by DNA compositional differences.…”

“…The difference in the response to the thermal exposure is obviously connected with the nature of these transitions. Their origin was recently considered by Staynov [5] and Bryan et al [12]. The following possibilities were suggested [ 5 ] : (a) the presence within each nucleosome of two regions with different thermal stability of DNA, (b) the existence of two structurally different classes of nucleosomes, and (c) the melting of nucleosomal DNA depends on the length of the adjacent spacer regions.…”

“…The question arises, therefore, about the relationship between the melting pattern of the polymer material and that of the monosomes. Recently, Bryan et al [12] forwarded the idea that the premelt and the main thermal transition of the monosomes corresponded to the 72 "C and 82 "C transitions of the polymer nucleoprotein, respectively, the transitions of the monomers being shifted to a lower temperature by the free DNA ends.In the present study on H1-depleted chromatin we developed an experimental approach which allowed us to obtain a material lacking the transition at 72 "C and to investigate some properties of this material as well as the melting profile of the monomer particles isolated from it. The data obtained support the hypothesis [I21 that the two high temperature transitons in HI -depleted chromatin reflect the presence within each nucleosome of two regions with different thermal stability of DNA.…”

“…The question arises, therefore, about the relationship between the melting pattern of the polymer material and that of the monosomes. Recently, Bryan et al [12] forwarded the idea that the premelt and the main thermal transition of the monosomes corresponded to the 72 "C and 82 "C transitions of the polymer nucleoprotein, respectively, the transitions of the monomers being shifted to a lower temperature by the free DNA ends.…”