title: Thermodynamic studies of the core histones: pH and ionic strength effects on the stability of the (H3-H4)/(H3-H4)2 system.
abstract: The self-associative behavior and the thermal stability of the H3/H4 histone complex was studied in low-ionic strength conditions by several physicochemical techniques, including differential scanning calorimetry and circular dichroism spectroscopy. At neutrality, the major molecular species present in solution is the (H3-H4)2 tetramer. Its thermodynamic properties cannot be studied directly though, since its thermal denaturation is completely irreversible even at the lowest salt concentrations. However, a complete thermodynamic analysis can be performed at low ionic strength and pH 4.5, where the (H3-H4)2 tetramer is quantitatively dissociated into two H3-H4 dimers and where almost complete reversibility of the thermal transitions is attained. The unfolding transition temperature of the 26.5 kDa H3-H4 dimer increases as a function of both the ionic strength of the solvent and the total protein concentration. The thermal denaturation of the H3-H4 dimer is characterized by the presence of a single calorimetric peak, centered at 58 degrees C, with a corresponding enthalpy change of 25 kcal/mol of a 13 kDa monomer unit and a change in heat capacity upon unfolding of about 0.6 kcal/(K mol of 13 kDa monomer unit). The complex between histones H3 and H4 (tetramer or dimer) is stable between pH 9.5 and 3.0. At pH 1.5, the system is almost completely unfolded at all temperatures. At low ionic strengths and pH values between 5.0 and 2.5, the H3-H4 dimer behaves as a highly cooperative system, melting as a single unit; i.e. individual H3 and H4 folded monomers are not detectable during the treatment. The two-state mechanism accounting for the unfolding of the H3-H4 dimer at pH 4.5 is the same as that described for the H2A-H2B dimer at neutrality. Just like for the H2A and H2B histones, the H3 and H4 polypeptides are properly folded only when assembled as H3-H4 dimers or in higher-order histone assemblies. Therefore, coupling along the interfaces of the two chains within the heterodimer is the major factor contributing to the stabilization of the secondary and tertiary structures of the chains as well as of the histone dimers.
authors.affiliation: Department of Biology, Johns Hopkins University, Baltimore, Maryland 21218, USA.
authors.forename: V
authors.initials: V
authors.lastname: Karantza
authors.forename: E
authors.initials: E
authors.lastname: Freire
authors.forename: E N
authors.initials: EN
authors.lastname: Moudrianakis
mesh_terms: D000818:Animals; D002152:Calorimetry, Differential Scanning; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002645:Chickens; D002942:Circular Dichroism; D004355:Drug Stability; D006657:Histones / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D009994:Osmolar Concentration; D011487:Protein Conformation; D011489:Protein Denaturation; D013816:Thermodynamics
chemical_list: D006657:Histones