Histone-DNA binding free energy cannot be measured in dilution-driven dissociation experiments.
Despite decades of study on nucleosomes, there has been no experimental determination of the free energy of association between histones and DNA. Instead, only the relative free energy of association of the histone octamer for differing DNA sequences has been available. Recently, a method was developed based on quantitative analysis of nucleosome dissociation in dilution experiments that provides a simple practical measure of nucleosome stability. Solution conditions were found in which nucleosome dissociation driven by dilution fit well to a simple model involving a noncooperative nucleosome assembly/disassembly equilibrium, suggesting that this approach might allow absolute equilibrium affinity of the histone octamer for DNA to be measured. Here, we show that the nucleosome assembly/disassembly process is not strictly reversible in these solution conditions, implying that equilibrium affinities cannot be obtained from these measurements. Increases in [NaCl] or temperature, commonly employed to suppress kinetic bottlenecks in nucleosome assembly, lead to cooperative behavior that cannot be interpreted with the simple assembly/disassembly equilibrium model. We conclude that the dilution experiments provide useful measures of kinetic but not equilibrium stability. Kinetic stability is of practical importance: it may govern nucleosome function in vivo, and it may (but need not) parallel absolute thermodynamic stability.
Thåström
A
A
Department of Biochemistry, Molecular Biology, and Cell Biology, Northwestern University, Evanston, Illinois 60208-3500, USA.
Gottesfeld
J M
JM
Luger
K
K
Widom
J
J
D000818:Animals; D002021:Buffers; D002645:Chickens; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D003902:Detergents; D005990:Glycerol; D006003:Glycogen; D006657:Histones / Q000737:chemistry* / Q000378:metabolism*; D007700:Kinetics; D009707:Nucleosomes / Q000737:chemistry / Q000378:metabolism; D017830:Octoxynol; D011092:Polyethylene Glycols; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D012996:Solutions; D013816:Thermodynamics; D014325:Tromethamine; D014982:Xenopus laevis
D002021:Buffers; D004268:DNA-Binding Proteins; D003902:Detergents; D006657:Histones; D009707:Nucleosomes; D021122:Protein Subunits; D012996:Solutions; D014325:Tromethamine; D011092:Polyethylene Glycols; D017830:Octoxynol; D006003:Glycogen; C010615:Nonidet P-40; D005990:Glycerol