Isolation of nonhistone-protein-rich chromatin fragments differing in composition and recovery between transcriptionally active and inactive sources.
Soluble chromatin was fragmented by deoxyribonuclease I[EC 3.1.4.5] or II[EC 3.1.4.6] digestion or sonication, followed by 0.7 m NaCl treatment, which preferentially removed DNA fragments and histones from the fragmented chromatin, but not from intact chromatin, leaving an insoluble fraction of DNA-histone-nonhistone protein complex  without appreciable redistribution of the chromatin components. Fragmentation of DNA and concentration of nonhistone proteins in the fraction were most effective with deoxyribonuclease I under the conditions used. Thus, only 1% of the DNA in transcriptionally inactive chicken erythrocyte chromatin was recovered, together with 2% of the total histone and 50-100% of the total nonhistone protein. Separate experiments showed that this DNA fraction was among the double-stranded segments having in average size of 210 base pairs and was associated with histone components responsible for the nucleosome conformation as well as most of the nonhistone protein components in the original chromatin. From transcriptionally active  chicken liver or calf thymus chromatin, however, 5-10% of the total DNA, 20-40% of the histone, and 80-100% of the nonhistone protein were recovered  in  the corresponding fraction. These results may reflect the native states and template activities in situ of various chromatins.
D000818:Animals; D002645:Chickens; D002843:Chromatin / Q000032:analysis*; D002868:Chromosomal Proteins, Non-Histone / Q000302:isolation & purification*; D004247:DNA / Q000032:analysis; D003851:Deoxyribonucleases; D004912:Erythrocytes / Q000032:analysis; D008099:Liver / Q000032:analysis; D013010:Sonication; D013950:Thymus Gland / Q000032:analysis; D014158:Transcription, Genetic
D002843:Chromatin; D002868:Chromosomal Proteins, Non-Histone; D004247:DNA; D003851:Deoxyribonucleases