title: In-cell thermodynamics and a new role for protein surfaces.
abstract: There is abundant, physiologically relevant knowledge about protein cores; they are hydrophobic, exquisitely well packed, and nearly all hydrogen bonds are satisfied. An equivalent understanding of protein surfaces has remained elusive because proteins are almost exclusively studied in vitro in simple aqueous solutions. Here, we establish the essential physiological roles played by protein surfaces by measuring the equilibrium thermodynamics and kinetics of protein folding in the complex environment of living Escherichia coli cells, and under physiologically relevant in vitro conditions. Fluorine NMR data on the 7-kDa globular N-terminal SH3 domain of Drosophila signal transduction protein drk (SH3) show that charge-charge interactions are fundamental to protein stability and folding kinetics in cells. Our results contradict predictions from accepted theories of macromolecular crowding and show that cosolutes commonly used to mimic the cellular interior do not yield physiologically relevant information. As such, we provide the foundation for a complete picture of protein chemistry in cells.
authors.affiliation: Department of Chemistry, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599;
authors.forename: Austin E
authors.initials: AE
authors.lastname: Smith
authors.affiliation: Department of Chemistry, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599;
authors.forename: Larry Z
authors.initials: LZ
authors.lastname: Zhou
authors.affiliation: Department of Chemistry, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599;
authors.forename: Annelise H
authors.initials: AH
authors.lastname: Gorensek
authors.affiliation: Department of Physical Chemistry II, Ruhr-Universität Bochum, 44780 Bochum, Germany;
authors.forename: Michael
authors.initials: M
authors.lastname: Senske
authors.affiliation: Department of Chemistry, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599; Department of Biochemistry and Biophysics, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599; Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599 gary_pielak@unc.edu.
authors.forename: Gary J
authors.initials: GJ
authors.lastname: Pielak
mesh_terms: D000818:Animals; D004330:Drosophila; D006860:Hydrogen Bonding; D009682:Magnetic Resonance Spectroscopy; D011506:Proteins / Q000737:chemistry*; D013499:Surface Properties; D013816:Thermodynamics
chemical_list: D011506:Proteins
keywords: in-cell NMR; protein NMR; protein folding; protein thermodynamics