title: Physical determinants of β-barrel membrane protein folding in lipid vesicles.
abstract: The spontaneous folding of two Neisseria outer membrane proteins, opacity-associated (Opa)(60) and Opa(50) into lipid vesicles was investigated by systematically varying bulk and membrane properties. Centrifugal fractionation coupled with sodium dodecyl sulfate polyacrylamide gel electrophoresis mobility assays enabled the discrimination of aggregate, unfolded membrane-associated, and folded membrane-inserted protein states as well as the influence of pH, ionic strength, membrane surface potential, lipid saturation, and urea on each. Protein aggregation was reduced with increasing lipid chain length, basic pH, low salt, the incorporation of negatively charged guest lipids, or by the addition of urea to the folding reaction. Insertion from the membrane-associated form was improved in shorter chain lipids, with more basic pH and low ionic strength; it is hindered by unsaturated or ether-linked lipids. The isolation of the physical determinants of insertion suggests that the membrane surface and dipole potentials are driving forces for outer membrane protein insertion and folding into lipid bilayers.
authors.affiliation: Department of Chemistry, University of Virginia, Charlottesville, Virginia, USA.
authors.forename: Alison H
authors.initials: AH
authors.lastname: Dewald
authors.forename: Jacqueline C
authors.initials: JC
authors.lastname: Hodges
authors.forename: Linda
authors.initials: L
authors.lastname: Columbus
mesh_terms: D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry* / Q000378:metabolism*; D002244:Carbon; D002498:Centrifugation; D005591:Chemical Fractionation; D004591:Electrophoresis, Polyacrylamide Gel; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D007700:Kinetics; D008051:Lipid Bilayers / Q000737:chemistry*; D008055:Lipids / Q000737:chemistry; D009994:Osmolar Concentration; D017510:Protein Folding / Q000187:drug effects; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D013499:Surface Properties / Q000187:drug effects; D014508:Urea / Q000494:pharmacology
chemical_list: D001425:Bacterial Outer Membrane Proteins; D005231:Fatty Acids, Unsaturated; D008051:Lipid Bilayers; D008055:Lipids; C078167:OMPA outer membrane proteins; C090348:Opa protein, Neisseria; D002244:Carbon; D014508:Urea