Reconstitution of native-like nucleosome core particles from reversed-phase-HPLC-fractionated histones

Abstract

We have reconstituted nucleosome core particles from reversed-phase-HPLC-purified chicken erythrocyte core histones and 145 bp random-sequence DNA fragments. Characterization of the resulting nucleoprotein complexes by sedimentation velocity, CD and DNase I footprinting showed that they are structurally indistinguishable from native nucleosome core particles. Furthermore, we have shown that the ability to reproduce these native-like structural features in these reconstituted nucleosomecore particles is basically independent of the biological source or the method used (i.e. salt versus acid) for the extraction of histones before their HPLC fractionation. The usefulness and relevance of this approach for the reconstitution of native-like chromatin structures from histone types (histone variants/post/translationally modified histones), which are usually available only in relatively small amounts, is discussed.