title: Gamma S-crystallin of bovine and human eye lens: solution structure, stability and folding of the intact two-domain protein and its separate domains.
abstract: Human and bovine gammaS-crystallin (HgammaS and BgammaS) and their isolated N- and C-terminal domains were cloned and expressed as recombinant proteins in E. coli. HgammaS and BgammaS are found to be authentic according to their spectral and hydrodynamic properties. Both full-length proteins and isolated domains are monomeric and exhibit high thermal and pH stabilities. The thermodynamic characterization made use of chemically and thermally-induced equilibrium unfolding transitions at varying pH. In spite of its exemplary two-domain structure, gammaS-crystallin does not show bimodal unfolding characteristics. In the case of BgammaS, at pH 7.0, the C-terminal domain is less stable than the N-terminal one, whereas for HgammaS the opposite holds true. Differential scanning calorimetry confirms the results of chemically-induced equilibrium unfolding transitions. Over the whole pH range between 2.0 and 11.5, HgammaS-crystallin and its isolated domains (HgammaS-N and HgammaS-C) follow the two-state model. The two-state unfolding of the intact two-domain protein points to the close similarity of the stabilities of the constituent domains. Obviously, interactions between the domains do not contribute significantly to the overall stability of gammaS-crystallin. In contrast, the structurally closely related gammaB-crystallin owes much of its extreme stability to domain interactions.
authors.affiliation: Institute of Biophysics and Physical Biochemistry, University of Regensburg, Germany.
authors.forename: M
authors.initials: M
authors.lastname: Wenk
authors.forename: R
authors.initials: R
authors.lastname: Herbst
authors.forename: D
authors.initials: D
authors.lastname: Hoeger
authors.forename: M
authors.initials: M
authors.lastname: Kretschmar
authors.forename: N H
authors.initials: NH
authors.lastname: Lubsen
authors.forename: R
authors.initials: R
authors.lastname: Jaenicke
mesh_terms: D000818:Animals; D002152:Calorimetry, Differential Scanning; D002417:Cattle; D002942:Circular Dichroism; D003001:Cloning, Molecular; D003459:Crystallins / Q000737:chemistry* / Q000235:genetics; D004229:Dithiothreitol / Q000494:pharmacology; D004926:Escherichia coli; D019791:Guanidine / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D007908:Lens, Crystalline / Q000737:chemistry*; D011489:Protein Denaturation / Q000187:drug effects; D017510:Protein Folding; D020673:Protein Renaturation / Q000187:drug effects; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D013696:Temperature; D013816:Thermodynamics
chemical_list: D003459:Crystallins; D011994:Recombinant Proteins; D019791:Guanidine; D004229:Dithiothreitol