Conformational changes induced by salt in complexes of histones and superhelical nuclear DNA.
When HeLa cells are lysed in solutions containing a non-ionic detergent and 0.75 M-NaCl, structures are released that retain many of the morphological features of nuclei. These nucleoids contain all the nuclear DNA, RNA and the 'core' histones, but few other proteins characteristic of chromatin. Their DNA is intact. The core histones dissociate on raising the salt concentration. We have probed the structure of nucleoid-histone complexes using the intercalating dye, ethidium, or the RNA polymerase of Escherichia coli. Both have a higher affinity for superhelical DNA than they do for relaxed DNA. The binding of ethidium is measured fluorometrically, and using this probe we find that the DNA of nucleoids containing all the core histones behaves as if it were supercoiled slightly positively. As the salt concentration is increased, free energy characteristic of negative supercoiling appears between 0.92 M and 0.95 M-NaCl. This transition, which is reversible in the presence of the arginine-rich histones, occurs without dissociation of these histones from the DNA and so must reflect a conformational change in the complex. In contrast to the results with ethidium, we find that RNA polymerase can detect the presence of some negative free energy of supercoiling in nucleoids containing the core histones. The transformations of the free energy that can assist the binding of ethidium and RNA polymerase are discussed.
D001665:Binding Sites / Q000187:drug effects; D004278:DNA, Superhelical / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D004996:Ethidium / Q000378:metabolism; D006367:HeLa Cells / Q000187:drug effects / Q000378:metabolism; D006657:Histones / Q000378:metabolism; D006801:Humans; D009690:Nucleic Acid Conformation / Q000187:drug effects; D012965:Sodium Chloride / Q000494:pharmacology; D013050:Spectrometry, Fluorescence
D004278:DNA, Superhelical; D006657:Histones; D012965:Sodium Chloride; D012321:DNA-Directed RNA Polymerases; D004996:Ethidium