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School of Molecular Biosciences, Washington State University, Pullman, Washington 99164-4660, USA
(RECEIVED July 30, 2004; FINAL REVISION October 5, 2004; ACCEPTED October 9, 2004)
The nucleosome, the basic fundamental repeating unit of chromatin, contains two H2A/H2B dimers and an H3/H4 tetramer. Modulation of the structure and dynamics of the nucleosome is an important regulation mechanism of DNA-based chemistries in the eukaryotic cell, such as transcription and replication. One means of altering the properties of the nucleosome is by incorporation of histone variants. To provide insights into how histone variants may impact the thermodynamics of the nucleosome, the stability of the heterodimer between the H2A.Z variant and H2B was determined by urea-induced denaturation, monitored by far-UV circular dichroism, intrinsic Tyr fluorescence intensity, and anisotropy. In the absence of stabilizing agents, the H2A.Z/H2B dimer is only partially folded. The stabilizing cosolute, trimethylamine-N-oxide (TMAO) was used to promote folding of the unstable heterodimer. The equilibrium stability of the H2A.Z/H2B dimer is compared to that of the H2A/H2B dimer. The equilibrium folding of both histone dimers is highly reversible and best described by a two-state model, with no detectable equilibrium intermediates populated. The free energies of unfolding, in the absence of denaturant, of H2A.Z/H2B and H2A/H2B are 7.3 kcal mol–1 and 15.5 kcal mol–1, respectively, in 1 M TMAO. The H2A.Z/H2B dimer is the least stable histone fold characterized to date, while H2A/H2B appears to be the most stable. It is speculated that this difference in stability may contribute to the different biophysical properties of nucleosomes containing the major H2A and the H2A.Z variant.
Keywords: thermodynamics; chemical denaturation; circular dichroism; fluorescence; histone variant
Abbreviations: CD, circular dichroism • FL, fluorescence • Fapp, apparent fraction of unfolded monomer • 
G° (H2O), the free energy of unfolding in the absence of denaturant • GdmCl, guanidinium chloride • H2A, major Xenopus laevis isoform • H2A.Z, H2A variant encoded by the mouse gene • H2A-C, C-terminal tail truncation, after Arg99, of the Xenopus laevis major H2A isoform • KPi, potassium phosphate m-value, parameter describing the sensitivity of the unfolding transition to [Urea] • MRE, mean residue ellipticity • NCP, nucleosome core particle • SVD, singular value decomposition • TMAO, trimethylamine-N-oxide
Article published online ahead of print. Article and publication date are at http://www.proteinscience.org/cgi/doi/10.1110/ps.041026405.
Reprint requests to: Lisa M. Gloss, School of Molecular Biosciences, Washington State University, Pullman, WA 99164-4660, USA; e-mail: lmgloss{at}wsu.edu; fax: (509) 335-9688.
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