Differential scanning calorimetry of nuclei reveals the loss of major structural features in chromatin by brief nuclease treatment.
Differential scanning calorimetry revealed that chromatin melts in four distinct transitions in intact HeLa nuclei at 60 degrees C, 76 degrees C, 88 degrees C, and 105 degrees C. Calorimetry of whole cells was characterized by the same four transitions along with another at 65 degrees C, which was probably RNA. Isolated chromatin, however, melted in only two transitions at 72 degrees C and 85 degrees C. Very brief digestion of HeLa nuclei with either micrococcal nuclease or DNase I resulted in the conversion of a structure that melted at 105 degrees C to one that melted at 88 degrees C. Further digestion with micrococcal nuclease to the level of the mononucleosome did not result in any further substantial changes in either enthalpy or melting temperatures. In contrast, further DNase I digestion that resulted in cleavage within the nucleosome produced a pronounced shift in melting temperatures to broad transitions at 62 degrees C and 78 degrees C.
D002152:Calorimetry, Differential Scanning; D002843:Chromatin / Q000187:drug effects* / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000494:pharmacology*; D006367:HeLa Cells / Q000378:metabolism; D006657:Histones / Q000378:metabolism; D006801:Humans; D008836:Micrococcal Nuclease / Q000494:pharmacology*; D009690:Nucleic Acid Conformation; D009707:Nucleosomes / Q000378:metabolism; D011487:Protein Conformation; D013816:Thermodynamics
D002843:Chromatin; D006657:Histones; D009707:Nucleosomes; D004247:DNA; D003850:Deoxyribonuclease I; D008836:Micrococcal Nuclease