The histone H3/H4.N1 complex supplemented with histone H2A-H2B dimers and DNA topoisomerase I forms nucleosomes on circular DNA under physiological conditions.
We have fractionated the whole cell extract of Xenopus oocytes (oocyte S-150) and isolated the endogenous components required for DNA supercoiling and nucleosome formation. Histone H2B and the three oocyte-specific H2A proteins were purified as free histones. Histones H3 and H4 were purified 100-fold in a complex with the acidic protein N1. In the presence of DNA topoisomerase I or II, histone H3/H4.N1 complexes supercoil DNA in a reaction that is inhibited by Mg2+, and this inhibition is relieved by NTPs. The supercoiling reaction induced by H3/H4.N1 complexes is enhanced by free histone H2A-H2B dimers, which by themselves do not supercoil DNA. Nuclease digestions and protein analyses indicate that H3/H4.N1 complexes form subnucleosomal particles containing histones H3 and H4. Nucleosomes containing 146-base pair DNA and the four histones are formed when histones H2A and H2B complement the reaction.
Zucker
K
K
Department of Biology, University of Rochester, New York.
Worcel
A
A
D000818:Animals; D002499:Centrifugation, Density Gradient; D002852:Chromatography, Ion Exchange; D004264:DNA Topoisomerases, Type I / Q000302:isolation & purification / Q000378:metabolism*; D004270:DNA, Circular / Q000302:isolation & purification / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004912:Erythrocytes / Q000378:metabolism; D005260:Female; D006657:Histones / Q000302:isolation & purification / Q000378:metabolism*; D046911:Macromolecular Substances; D008836:Micrococcal Nuclease; D008970:Molecular Weight; D009687:Nuclear Proteins / Q000302:isolation & purification / Q000378:metabolism; D056489:Nucleoplasmins; D009707:Nucleosomes / Q000378:metabolism* / Q000648:ultrastructure; D009865:Oocytes / Q000378:metabolism; D010750:Phosphoproteins; D014982:Xenopus laevis
D004270:DNA, Circular; D006657:Histones; D046911:Macromolecular Substances; D009687:Nuclear Proteins; D056489:Nucleoplasmins; D009707:Nucleosomes; D010750:Phosphoproteins; D008836:Micrococcal Nuclease; D004264:DNA Topoisomerases, Type I