Quantitative analysis of DNase I digestion patterns of oligo- and polynucleosomes

Journal of Molecular Biology
D Z Staynov, Y G Proykova

Abstract

DNase I digestion of unlabelled chromatin has been used to determine the orientation of nucleosomes in the higher-order structure of chromatin. DNA digestion patterns were analysed quantitatively and compared with simulation curves that were generated with the experimentally obtained rate constants for cutting inside nucleosomes and the half-height widths of the bands. The rate constants for cutting at the linker DNA were varied to fit the experimental curves. By comparing the digestion profiles of polynucleosomes, oligonucleosomes and H1/H5-depleted oligonucleosomes we have been able to distinguish between protection caused by H1/H5 histones only and protection caused by the higher-order structure. By the nature of this method the area protected by H1/H5 histones appears symmetrical around the centre of the nucleosomal DNA on the dyad axis (position S[0]), but it can also be interpreted as a superposition of two separate protected areas that are symmetrical around position S[0]. Protection by the higher-order structure shows that the nucleosomes are oriented with their S[0] positions inside the 30 nm fibre and that there is a minimum number of nucleosomes required for this structure to be formed. We have also found that the li...Continue Reading

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Citations

May 23, 2017·Nature Communications·Jelka PoharMojca Benčina
Jan 12, 1999·FEBS Letters·S Belikov, V Karpov
Jun 13, 2009·Chromosome Research : an International Journal on the Molecular, Supramolecular and Evolutionary Aspects of Chromosome Biology·Samantha SantangeloDontcho Z Staynov
May 15, 2004·British Journal of Haematology·K B PetersenC Guldborg Nyvold

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