A topoisomerase from chicken erythrocyte nuclei which does not assemble nucleosome core particles in vitro.
We have examined the ability of a topoisomerase purified from chicken erythrocyte nuclei to mediate nucleosome core assembly in vitro at physiological ionic strength (0.15 M NaCl). Although we have detected limited amounts of spontaneously assembled nucleosome cores at this salt concentration, the addition of this topoisomerase does not increase the amount of assembly observed. Nucleosome assembly was assayed by quantitating the amount of core particle length DNA accumulated with time upon the nuclease digestion of histone-DNA complexes. In addition, the amount of negative supercoils introduced into relaxed closed circular DNA upon nucleosome core particle assembly was determined. Correctly assembled complexes do not protect more DNA from nuclease digestion than random histone-DNA complexes but shift the heterogeneous size distribution of protected fragments to a more homogeneous distribution centred around 145 base pairs. Under our conditions of nucleosome assembly, a second histone-DNA complex which is distinct from the nucleosome core can be detected under physiological ionic strength conditions. This particle does not form in high salt assembly experiments. Similarly, the assembly of this particle is unaffected by the presence or absence of topoisomerase.
D000818:Animals; D002467:Cell Nucleus / Q000201:enzymology*; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002645:Chickens; D004247:DNA / Q000096:biosynthesis; D004264:DNA Topoisomerases, Type I / Q000097:blood / Q000302:isolation & purification / Q000378:metabolism*; D004912:Erythrocytes / Q000201:enzymology*; D006657:Histones / Q000302:isolation & purification; D006868:Hydrolysis; D066298:In Vitro Techniques; D009707:Nucleosomes / Q000378:metabolism*; D012995:Solubility
D006657:Histones; D009707:Nucleosomes; D004247:DNA; D004264:DNA Topoisomerases, Type I