Characterization of the histone core complex.

Abstract

A stable histone core complex containing equimolar ratios of H2A, H2B, H3, and H4 has been isolated from chicken erythrocyte chromatin in high salt. This complex has been characterized by sedimentation and chemical crosslinking studies. In velocity ultracentrifugation, only one single sharp sedimentation boundary has been observed with sedimentation coefficient S = 3.8 L 0.1. The molecular weight of the histone core com has been investigated by low-speed sedimentation equilibrium studies over a wide range of protein concentration. Analysis of the apparent weight-average molecular weight as a function of concentration indicates that the histone core complex in 2 M NaCl, pH 9.0, is in equilibrium between a tetramer (H2AXH2BXH3XH4) and an octamer I(H2AXH2BXH3XH4)12 species with a tetramer molecular weight of 55000. The equilibrium constant K is approximately 1.2 X 10-5 liter per mol at 100. Evidence of such a tetramer-octamer equilibrium in solution is also supported by the results of our chemical crosslinking experiments on the histone core complex. A well-characterized entity within the basic subunit of chromatin, the nucleosome, is the "core particle," which has a molecular weight of 200,000 and contains approximately 140 base pairs of DNA and two molecules each of the histones H2A, H2B, H3, and H4 (1, 2). Electron microscopy (3, 4), neutron scattering (5, 6), and x-ray diffraction (7) studies indicate that the "core particle" is a disk 110 A wide by 55 A high with two turns of DNA localized around the compact histone center. The arrangement of histones in the center and along the DNA within a nucleosome is not known. Proximity of certain histones within the nucleosome has been established by protein crosslinking studies; in particular, the use of zero-length crosslinking agents, ultraviolet light, tetranitromethane, and carbodiimide, has demonstrated close contact between H2B and H4, H2B and H2A, and H3 and H4, respectively (8, 9); furthermore, H2B is probably bound simultaneously to both H2A and H4 in vvo (8). Solution studies on mixtures of isolated histones have shown that histones H2A, H2B, H3, and H4 interact pairwise and that each possesses at least two distinct domains for interaction with other histones (9). Their interactions can be roughly divided into two categories: strong interactions between the pairs H2A-H2B, H2B-H4, and H3-H4, and weak interactions between the pairs H2A-H3, H2B-H3, and H2A-H4 (9). Along with dimeric complexes found in mixtures of histone pairs, the arginine-rich histones form a tetramer [(H3)(H4)]2 (10-13). In addition, more complex oligomeric structures are found in solutions of high ionic strength (2 M NaCl) at neutral to alkaline pH (pH 7-10) after gentle extraction of the chromatin with high salt, although the precise nature of the oligomers is not clear (6,(14)(15)(16)(17). The elevated ionic strength and pH reduce the repulsions between the highly charged histones, possibly providing an environment similar to that provided by DNA in the nucleosome. In fact, the c...

Reconstitution of native-like nucleosome core particles from reversed-phase-HPLC-fractionated histones

“…Under the experimental conditions (see Figure 5A) native core histones sediment as a monodisperse sample, with 3.8p0.2 S, in agreement with previous reports [28][29][30][31]. In contrast, RP-HPLCpurified histones (fraction C, Figure 1) exhibit a significant extent of heterogeneity ( Figure 5A2), with only approx.…”

Acetylation Mimics within Individual Core Histone Tail Domains Indicate Distinct Roles in Regulating the Stability of Higher-Order Chromatin Structure

“…H2A and H2B supernatants were mixed together and purified as described previously (16). The quality of dimers and tetramers was analyzed by 15% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and concentrations were determined using an A 230 of 4.2 for 1 mg/ml (6).…”

Nucleosome core histone complex isolated gently and rapidly in 2 M NaCl is octameric.

“…Two heterotypic tetramers are envisaged to be paired to form the nucleosome core protein (3,6,14). Chung et al (15) presented hydrodynamic and chemical data on the histone core complex in 2 M NaCl that support such an association of heterotypic tetramers to form the octamer.…”