title: Structure and stability of histone HMf from the hyperthermophilic archaeon Methanothermus fervidus.
abstract: The secondary and quaternary structures and stabilities of recombinant (r) forms of the HMfA and HMfB histones from Methanothermus fervidus have been investigated by CD spectroscopy and formaldehyde-mediated protein-protein cross-linking. Both proteins were shown to be dimers in solutions containing 5-1300 mM KCl, at pH 6-10 and 25-83 degrees C, and specifically in 1 M KCl, at pH 7.5 and 83 degrees C, conditions which approximate those in vivo in M. fervidus cells. Heat treatment of a mixture of rHMfA and rHMfB homodimers resulted in the formation of rHMfA.rHMfB heterodimers, as demonstrated by two-dimensional PAGE. Heterodimer formation did not result in a CD-detectable conformational change from the homodimer states, indicating that homogeneous (rHMfA)2 and (rHMfB)2 preparations may be considered as structural models of heterodimers. At pH 2, both rHMfA and rHMfB were denatured under low-salt (< 0.2 M KCl) conditions, and their conformations were stabilized in a cooperative manner by increasing KCl concentration, with cooperativity constants for KCl uptake of 2.7 and 3.1, respectively. The alpha-helical conformations of rHMfA and rHMfB were salt-dependent, at both pH 2 and pH 7.5, with maximal helicities in 1 M KCl of 84% and 63% at pH 2, and 72% and 65% at pH 7.5, respectively. The data obtained indicate that the structures of HMfA and HMfB, in 100-200 mM KCl at pH 7.5 and 25 degrees C, are likely to be very similar to their in vivo structures, even though these conditions are far removed from those found in vivo.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Microbiology, Ohio State University, Columbus 43210, USA.
authors.forename: R A
authors.initials: RA
authors.lastname: Grayling
authors.forename: W J
authors.initials: WJ
authors.lastname: Becktel
authors.forename: J N
authors.initials: JN
authors.lastname: Reeve
mesh_terms: D000595:Amino Acid Sequence; D002942:Circular Dichroism; D003432:Cross-Linking Reagents; D004355:Drug Stability; D004591:Electrophoresis, Polyacrylamide Gel; D019605:Euryarchaeota / Q000378:metabolism*; D005557:Formaldehyde; D006657:Histones / Q000737:chemistry* / Q000302:isolation & purification; D006358:Hot Temperature; D008969:Molecular Sequence Data; D009994:Osmolar Concentration; D011189:Potassium Chloride / Q000378:metabolism; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification; D017386:Sequence Homology, Amino Acid
chemical_list: D003432:Cross-Linking Reagents; D006657:Histones; D011994:Recombinant Proteins; C094495:histone HMf; D005557:Formaldehyde; D011189:Potassium Chloride