title: Structural stability and solution structure of chaperonin GroES heptamer studied by synchrotron small-angle X-ray scattering.
abstract: The GroES protein from Escherichia coli is a well-known member of the molecular chaperones. GroES consists of seven identical 10 kDa subunits, and forms a dome-like oligomeric structure. In order to obtain information on the structural stability and unfolding-refolding mechanism of GroES protein, especially at protein concentrations (0.4-1.2 mM GroES monomer) that would mimic heat stress conditions in vivo, we have performed synchrotron small-angle X-ray scattering (SAXS) experiments. Surprisingly, in spite of the high protein concentration, reversibility in the unfolding-refolding reaction was confirmed by SAXS experiments structurally. Although the unfolding-refolding reaction showed an apparent single transition with a Cm of 1.1 M guanidium hydrochloride, a more detailed analysis of this transition demonstrated that the unfolding mechanism could be best explained by a sequential three-state model, which consists of native heptamer, dissociated monomer, and unfolded monomer. Together with our previous result that GroES unfolded completely via a partially folded monomer according to a three-state model at low protein concentration (5 microM monomer), the unfolding-refolding mechanism of GroES protein could be explained uniformly by the three-state model from low to high protein concentrations. Furthermore, to clarify an ambiguity of the native GroES structure in solution, especially mobile loop structures, we have estimated a solution structure of GroES using SAXS profiles obtained from experiments and simulation analysis. The result suggested that the native structure of GroES in solution was very similar to that seen in GroES-GroEL complex determined by crystallography.
authors.affiliation: Department of Biotechnology, Faculty of Engineering, Tottori University, Koyama-Minami, Tottori 680-8552, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Higurashi
authors.forename: Yuzuru
authors.initials: Y
authors.lastname: Hiragi
authors.forename: Kaoru
authors.initials: K
authors.lastname: Ichimura
authors.forename: Yasutaka
authors.initials: Y
authors.lastname: Seki
authors.forename: Kunitsugu
authors.initials: K
authors.lastname: Soda
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Mizobata
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Kawata
mesh_terms: D018835:Chaperonin 10 / Q000737:chemistry*; D002850:Chromatography, Gel; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000737:chemistry; D029968:Escherichia coli Proteins / Q000737:chemistry*; D008958:Models, Molecular; D017510:Protein Folding; D020673:Protein Renaturation; D020836:Protein Structure, Quaternary; D021122:Protein Subunits; D012996:Solutions / Q000737:chemistry; D013329:Structure-Activity Relationship; D017356:Synchrotrons; D013816:Thermodynamics
chemical_list: D018835:Chaperonin 10; D029968:Escherichia coli Proteins; D021122:Protein Subunits; D012996:Solutions