Asymmetric Nucleosome Disassembly with Disrupted Histones Revealed by Time Resolved Small Angle X-Ray Scattering with Contrast Variation

Abstract

DNA packages into compact chromatin structure in eukaryotic cell despite its high negative charge. Nucleosome core particles (NCP) are the fundamental repeating units of chromatin and modulate DNA accessibility for gene expression and regulation. NCPs contain a symmetric histone octamer wrapped by 147-bp DNA. The dynamics of DNA packaging and unpackaging from NCPs affects all DNA-based chemistries, but is not well understood due to a lack of structures of the partially unwrapped, kinetic intermediates. Here we applied a novel strategy combining contrast variation with time-resolved small angle x-ray scattering to determine the structures of protein and DNA constituents of NCPs during salt-induced disassembly. We monitored DNA conformation and protein dissociation of NCPs with two positioning sequences: Widom 601 and 5S DNA. For the Widom 601 construct, we measure a transient structure on the millisecond time scale where the DNA is asymmetrically released from a disrupted histone core, and the proteins remain bound to unwrapped DNA in a semi-open conformation. We hypothesis this conformation may be biologically important substrate for gene regulation. The 5S construct also displays asymmetric DNA release, but exhibits a different pattern of protein dissociation. Our results establish a powerful platform for studying the global dynamics of nucleoprotein complexes.