Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

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Record number 360773
Title Tertiary structure in 7.9 M guanidinium chloride: the role of Glu-53 and Asp-287 in Pyrococcus furiosus endo-beta-1,3-glucanase
Author(s) Chiaraluce, R.; Florio, R.; Angelaccio, S.; Gianese, G.; Lieshout, J.F.T. van; Oost, J. van der; Consalvi, V.
Source FEBS Journal 274 (2007). - ISSN 1742-464X - p. 6167 - 6179.
DOI https://doi.org/10.1111/j.1742-4658.2007.06137.x
Department(s) Microbiological Laboratory
VLAG
Publication type Refereed Article in a scientific journal
Publication year 2007
Keyword(s) denatured state - staphylococcal nuclease - protein stability - unfolded state - thermophilic proteins - residual structure - m-values - intermediate - thermodynamics - salt
Abstract The thermodynamic stability of family 16 endo-ß-1,3-glucanase (EC 3.2.1.39) from the hyperthermophilic archaeon Pyrococcus furiosus is decreased upon single (D287A, E53A) and double (E53A/D287A) mutation of Asp287 and Glu53. In accordance with the homology model prediction, both carboxylic acids are involved in the composition of a calcium binding site, as shown by titration of the wild-type and the variant proteins with a chromophoric chelator. The present study shows that, in P. furiosus, endo-ß-1,3-glucanase residues Glu53 and Asp287 also make up a calcium binding site in 7.9 m guanidinium chloride. The persistence of tertiary structure in 7.9 m guanidinium chloride, a feature of the wild-type enzyme, is observed also for the three variant proteins. The ¿GH2O values relative to the guanidinium chloride-induced equilibrium unfolding of the three variants are approximatelty 50% lower than that of the wild-type. The destabilizing effect of the combined mutations of the double mutant is non-additive, with an energy of interaction of 24.2 kJ·mol¿1, suggesting a communication between the two mutated residues. The decrease in the thermodynamic stability of D287A, E53A and E53A/D287A is contained almost exclusively in the m-values, a parameter which reflects the solvent-exposed surface area upon unfolding. The decrease in m-value suggests that the substitution with alanine of two evenly charged repulsive side chains induces a stabilization of the non-native state in 7.9 m guanidinium chloride comparable to that induced by the presence of calcium on the wild-type. These results suggest that the stabilization of a compact non-native state may be a strategy for P. furiosus endo-ß-1,3-glucanase to thrive under adverse environmental conditions.
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