title: Polar interactions trump hydrophobicity in stabilizing the self-inserting membrane protein Mistic.
abstract: Canonical integral membrane proteins are attached to lipid bilayers through hydrophobic transmembrane helices, whose topogenesis requires sophisticated insertion machineries. By contrast, membrane proteins that, for evolutionary or functional reasons, cannot rely on these machineries need to resort to driving forces other than hydrophobicity. A striking example is the self-inserting Bacillus subtilis protein Mistic, which is involved in biofilm formation and has found application as a fusion tag supporting the recombinant production and bilayer insertion of other membrane proteins. Although this unusual protein contains numerous polar and charged residues and lacks characteristic membrane-interaction motifs, it is tightly bound to membranes in vivo and membrane-mimetic systems in vitro. Therefore, we set out to quantify the contributions from polar and nonpolar interactions to the coupled folding and insertion of Mistic. To this end, we defined conditions under which the protein can be unfolded completely and reversibly from various detergent micelles by urea in a two-state equilibrium and where the unfolded state is independent of the detergent used for solubilizing the folded state. This enabled equilibrium unfolding experiments previously used for soluble and β-barrel membrane proteins, revealing that polar interactions with ionic and zwitterionic headgroups and, presumably, the interfacial dipole potential stabilize the protein much more efficiently than nonpolar interactions with the micelle core. These findings unveil the forces that allow a protein to tightly interact with a membrane-mimetic environment without major hydrophobic contributions and rationalize the differential suitability of detergents for the extraction and solubilization of Mistic-tagged membrane proteins.
authors.affiliation: Molecular Biophysics, University of Kaiserslautern , Erwin-Schrödinger-Straβe 13, 67663 Kaiserslautern, Germany.
authors.forename: Jana
authors.initials: J
authors.lastname: Broecker
authors.forename: Sebastian
authors.initials: S
authors.lastname: Fiedler
authors.forename: Katharina
authors.initials: K
authors.lastname: Gimpl
authors.forename: Sandro
authors.initials: S
authors.lastname: Keller
mesh_terms: D001412:Bacillus subtilis / Q000737:chemistry*; D001426:Bacterial Proteins / Q000737:chemistry*; D057927:Hydrophobic and Hydrophilic Interactions; D008051:Lipid Bilayers / Q000737:chemistry; D008565:Membrane Proteins / Q000737:chemistry*; D055550:Protein Stability; D058767:Protein Unfolding
chemical_list: D001426:Bacterial Proteins; D008051:Lipid Bilayers; D008565:Membrane Proteins