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The assembly of an H2A2,H2B2,H3,H4 hexamer onto DNA under conditions of physiological ionic strength.

1983; Elsevier BV; Volume: 258; Issue: 21 Linguagem: Inglês

10.1016/s0021-9258(17)44117-2

1083-351X

Michael J. Ellison, David E. Pulleyblank,

DNA and Nucleic Acid Chemistry

A novel nucleohistone particle is generated in high yield when a complex of DNA with the four core histones formed under conditions that are close to physiological (0.15 M NaC1, pH 8) is treated with micrococcal nuclease.The particle was found to contain 102 base pairs of DNA in association with six molecules of histones in the ratio 2H2A:2H2B:lH3:1H4 after relatively brief nuclease treatment.Prolonged nuclease digestion resulted in a reduction in the DNA length to a sharply defined 92-base pair fragment that was resistant to further degradation.Apparently normal nucleosome core particles containing two molecules each of the four core histones in association with 145 base pairs of DNA and a particle containing one molecule each of histones H2A and H2B in association with approximately 40 base pairs of DNA were also generated during nuclease treatment of the histone-DNA complexes formed under physiological ionic strength conditons.Kinetic studies have shown that the hexamer particle is not a subnucleosomal fragment produced by the degradation of nucleosome core particles.Furthermore, the hexamer particle was not found among the products of nuclease digestion when histones and DNA were previously assembled in 0.6 M NaCl.The high sedimentation coefficient of the hexameric complex (8 S) suggests that the DNA component of the particle has a folded conformation.Although the structure of the nucleosome core has been extensively characterized (reviewed in Ref. l), there is little information on the range of structural alternatives which subunits of the nucleosome are able to adopt in response to transcriptional or replicational processes.Several lines of evidence indicate that the nucleosome core is composed of two types of operative subunit: the (H3,H4), tetramer and two (H2A,H2B) pairs (2-12).Most of the structural integrity of the nucleosome core has been attributed to the binding of an (H3,H4), tetramer to the DNA ( 5 , 8, 13-21), while the (H2A,H2B) pairs have been considered to have a more passive function in the organization of the nucleosome (12, 18).Studies of the mechanism of histone assembly into nucleosome cores have also supported the view that the (H3,H4), tetramer and the (H2A,H2B) pair represent discrete substructures of the nucleosome core.Ruiz-Carrillo and co-workers (21, 23) have reported that the (H3,H4), tetramer is a key