title: Mapping the energy landscape for second-stage folding of a single membrane protein.
abstract: Membrane proteins are designed to fold and function in a lipid membrane, yet folding experiments within a native membrane environment are challenging to design. Here we show that single-molecule forced unfolding experiments can be adapted to study helical membrane protein folding under native-like bicelle conditions. Applying force using magnetic tweezers, we find that a transmembrane helix protein, Escherichia coli rhomboid protease GlpG, unfolds in a highly cooperative manner, largely unraveling as one physical unit in response to mechanical tension above 25 pN. Considerable hysteresis is observed, with refolding occurring only at forces below 5 pN. Characterizing the energy landscape reveals only modest thermodynamic stability (ΔG = 6.5 kBT) but a large unfolding barrier (21.3 kBT) that can maintain the protein in a folded state for long periods of time (t1/2 ∼3.5 h). The observed energy landscape may have evolved to limit the existence of troublesome partially unfolded states and impart rigidity to the structure.
authors.affiliation: National Creative Research Initiative Center for Single-Molecule Systems Biology, KAIST, Daejeon, South Korea.|Department of Physics, KAIST, Daejeon, South Korea.
authors.forename: Duyoung
authors.initials: D
authors.lastname: Min
authors.affiliation: Department of Chemistry and Biochemistry, University of California-Los Angeles, Los Angeles, California, USA.
authors.forename: Robert E
authors.initials: RE
authors.lastname: Jefferson
authors.affiliation: Department of Chemistry and Biochemistry, University of California-Los Angeles, Los Angeles, California, USA.
authors.forename: James U
authors.initials: JU
authors.lastname: Bowie
authors.affiliation: National Creative Research Initiative Center for Single-Molecule Systems Biology, KAIST, Daejeon, South Korea.|Department of Physics, KAIST, Daejeon, South Korea.
authors.forename: Tae-Young
authors.initials: TY
authors.lastname: Yoon
mesh_terms: D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism; D007700:Kinetics; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D011487:Protein Conformation; D017510:Protein Folding; D013816:Thermodynamics
chemical_list: D029968:Escherichia coli Proteins; D008565:Membrane Proteins