title: Out but not in: the large transmembrane β-barrel protein FhuA unfolds but cannot refold via β-hairpins.
abstract: How transmembrane β-barrel proteins insert and fold into membranes and by which factors they destabilize, unfold, and misfold represents a field of intense studies. Here, we use single-molecule force spectroscopy to characterize the un- and refolding of the ferric hydroxamate uptake receptor (FhuA), which is one of the largest β-barrel proteins of the outer membrane of Escherichia coli. Applied to mechanical stress, FhuA undergoes a complex unfolding pathway in which each of the 11 β-hairpins unfolds one after the other until the entire β-barrel has unfolded. Once unfolded and relaxed, the FhuA polypeptide cannot fold back into the lipid membrane and adopts various misfolded conformations. Such misfolding is in contrast to the reversible refolding behavior of much smaller β-barrel outer membrane proteins OmpA and OmpG that occurs at similar experimental conditions. The results suggest that large β-barrel proteins that show more complex (un-)folding pathways require cofactors for proper insertion and folding into the membrane.
authors.affiliation: Biosystems Science and Engineering (BSSE), ETH Zürich, Mattenstrasse 26, 4058 Basel, Switzerland.
authors.forename: Johannes
authors.initials: J
authors.lastname: Thoma
authors.forename: Patrick
authors.initials: P
authors.lastname: Bosshart
authors.forename: Moritz
authors.initials: M
authors.lastname: Pfreundschuh
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Müller
mesh_terms: D020816:Amino Acid Motifs; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry* / Q000648:ultrastructure; D004926:Escherichia coli; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000648:ultrastructure; D005583:Fourier Analysis; D008051:Lipid Bilayers / Q000737:chemistry; D018625:Microscopy, Atomic Force; D008958:Models, Molecular; D058849:Protein Refolding; D055550:Protein Stability; D017433:Protein Structure, Secondary; D058767:Protein Unfolding
chemical_list: D001425:Bacterial Outer Membrane Proteins; D029968:Escherichia coli Proteins; C050900:FhuA protein, E coli; D008051:Lipid Bilayers