The role of membrane destabilisation and protein dynamics in BAM catalysed OMP folding

DOI: 10.1038/s41467-021-24432-x

Authors: Paul White (University of Leeds) , Samuel F. Haysom (University of Leeds) , Matthew G. Iadanza (University of Leeds) , Anna J. Higgins (University of Leeds) , Jonathan M. Machin (University of Leeds) , James M. Whitehouse (University of Leeds) , Jim E. Horne (University of Leeds) , Bob Schiffrin (University of Leeds) , Charlotte Carpenter-Platt (University of Leeds) , Antonio N. Calabrese (University of Leeds) , Kelly M. Storek (Genentech Inc) , Steven T. Rutherford (Genentech Inc) , David J. Brockwell (University of Leeds) , Neil A. Ranson (University of Leeds) , Sheena E. Radford (University of Leeds)
Co-authored by industrial partner: Yes

Type: Journal Paper
Journal: Nature Communications , VOL 12

State: Published (Approved)
Published: July 2021
Diamond Proposal Number(s): 19248

Abstract: The folding of β-barrel outer membrane proteins (OMPs) in Gram-negative bacteria is catalysed by the β-barrel assembly machinery (BAM). How lateral opening in the β-barrel of the major subunit BamA assists in OMP folding, and the contribution of membrane disruption to BAM catalysis remain unresolved. Here, we use an anti-BamA monoclonal antibody fragment (Fab1) and two disulphide-crosslinked BAM variants (lid-locked (LL), and POTRA-5-locked (P5L)) to dissect these roles. Despite being lethal in vivo, we show that all complexes catalyse folding in vitro, albeit less efficiently than wild-type BAM. CryoEM reveals that while Fab1 and BAM-P5L trap an open-barrel state, BAM-LL contains a mixture of closed and contorted, partially-open structures. Finally, all three complexes globally destabilise the lipid bilayer, while BamA does not, revealing that the BAM lipoproteins are required for this function. Together the results provide insights into the role of BAM structure and lipid dynamics in OMP folding.

Journal Keywords: Cellular microbiology; Cryoelectron microscopy; Membrane structure and assembly; Protein folding

Diamond Keywords: Bacteria

Subject Areas: Biology and Bio-materials, Chemistry


Instruments: I24-Microfocus Macromolecular Crystallography

Added On: 26/07/2021 14:00

Documents:
s41467-021-24432-x.pdf

Discipline Tags:

Biochemistry Catalysis Chemistry Structural biology Life Sciences & Biotech

Technical Tags:

Diffraction Macromolecular Crystallography (MX)