title: Boosting protein stability with the computational design of β-sheet surfaces.
abstract: β-sheets often have one face packed against the core of the protein and the other facing solvent. Mutational studies have indicated that the solvent-facing residues can contribute significantly to protein stability, and that the preferred amino acid at each sequence position is dependent on the precise structure of the protein backbone and the identity of the neighboring amino acids. This suggests that the most advantageous methods for designing β-sheet surfaces will be approaches that take into account the multiple energetic factors at play including side chain rotamer preferences, van der Waals forces, electrostatics, and desolvation effects. Here, we show that the protein design software Rosetta, which models these energetic factors, can be used to dramatically increase protein stability by optimizing interactions on the surfaces of small β-sheet proteins. Two design variants of the β-sandwich protein from tenascin were made with 7 and 14 mutations respectively on its β-sheet surfaces. These changes raised the thermal midpoint for unfolding from 45°C to 64°C and 74°C. Additionally, we tested an empirical approach based on increasing the number of potential salt bridges on the surfaces of the β-sheets. This was not a robust strategy for increasing stability, as three of the four variants tested were unfolded.
authors.affiliation: Department of Biochemistry and Biophysics, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina.
authors.forename: Doo Nam
authors.initials: DN
authors.lastname: Kim
authors.affiliation: Program in Bioinformatics and Computational Biology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina.
authors.forename: Timothy M
authors.initials: TM
authors.lastname: Jacobs
authors.affiliation: Department of Biochemistry and Biophysics, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina.|Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina.
authors.forename: Brian
authors.initials: B
authors.lastname: Kuhlman
mesh_terms: D018360:Crystallography, X-Ray; D006801:Humans; D008958:Models, Molecular; D011487:Protein Conformation; D055550:Protein Stability; D017433:Protein Structure, Secondary; D058767:Protein Unfolding; D012984:Software; D019063:Tenascin / Q000737:chemistry*; D013816:Thermodynamics
chemical_list: D019063:Tenascin
keywords: Rosetta molecular modeling program; charge zipper proteins; computational protein design; electrostatic interactions; protein stability; β-sheets