Biophysical characterization of the centromere-specific nucleosome from budding yeast

Abstract

The centromeric DNA of all eukaryotes is assembled upon a specialized nucleosome containing a histone H3 variant known as CenH3. Despite the importance and conserved nature of this protein, the characteristics of the centromeric nucleosome are still poorly understood. In particular, the stoichiometry and DNA-binding properties of the CenH3 nucleosome have been the subject of some debate. We have characterized the budding yeast centromeric nucleosome by biochemical and biophysical methods and show that it forms a stable octamer containing two copies of the Cse4 protein and wraps DNA in a left-handed supercoil, similar to the canonical H3 nucleosome. The DNA-binding properties of the recombinant nucleosome are identical to those observed in vivo demonstrating that the octameric structure is physiologically relevant. © 2011 by The American Society for Biochemistry and Molecular Biology, Inc.