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Genetic Engineering of an Unconventional Yeast for Renewable Biofuel and Biochemical Production
Published on: September 20, 2016
Biochemical Analysis of Dimethyl Suberimidate-crosslinked Yeast Nucleosomes
Yuichi Ichiakwa1, Paul D Kaufman1
1Department of Molecular, Cell, and Cancer Biology, University of Massachusetts Medical School, Worcester, MA, USA.
None:
Nucleosomes are the fundamental unit of eukaryotic chromosome packaging, comprised of 147 bp of DNA wrapped around two molecules of each of the core histone proteins H2A, H2B, H3, and H4. Nucleosomes are symmetrical, with one axis of symmetry centered on the homodimeric interaction between the C-termini of the H3 molecules. To explore the functional consequences of nucleosome symmetry, we designed an obligate pair of H3 heterodimers, termed H3X and H3Y, allowing us to compare cells with single or double H3 alterations. Our biochemical validation of the heterodimeric X-Y interaction included intra-nucleosomal H3 crosslinking using dimethyl suberimidate (DMS). Here, we provide a detailed protocol for the use of DMS to analyze yeast nucleosomes.
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