Outer-membrane translocation of bulky small molecules by passive diffusion

Proc Natl Acad Sci U S A. 2015 Jun 9;112(23):E2991-9. doi: 10.1073/pnas.1424835112. Epub 2015 May 26.

Abstract

The outer membrane (OM) of gram-negative bacteria forms a protective layer around the cell that serves as a permeability barrier to prevent unrestricted access of noxious substances. The permeability barrier of the OM results partly from the limited pore diameters of OM diffusion channels. As a consequence, there is an "OM size-exclusion limit," and the uptake of bulky molecules with molecular masses of more than ∼ 600 Da is thought to be mediated by TonB-dependent, active transporters. Intriguingly, the OM protein CymA from Klebsiella oxytoca does not depend on TonB but nevertheless mediates efficient OM passage of cyclodextrins with diameters of up to ∼ 15 Å. Here we show, by using X-ray crystallography, molecular dynamics simulations, and single-channel electrophysiology, that CymA forms a monomeric 14-stranded β-barrel with a large pore that is occluded on the periplasmic side by the N-terminal 15 residues of the protein. Representing a previously unidentified paradigm in OM transport, CymA mediates the passive diffusion of bulky molecules via an elegant transport mechanism in which a mobile element formed by the N terminus acts as a ligand-expelled gate to preserve the permeability barrier of the OM.

Keywords: CymA; cyclodextrin; ligand gating; outer membrane channel; passive diffusion.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacterial Outer Membrane Proteins / metabolism
  • Binding Sites
  • Biological Transport*
  • Cell Membrane / metabolism
  • Crystallography, X-Ray
  • Cyclodextrins / metabolism
  • Diffusion
  • Klebsiella oxytoca / metabolism*
  • Molecular Dynamics Simulation

Substances

  • Bacterial Outer Membrane Proteins
  • Cyclodextrins

Associated data

  • PDB/4D51
  • PDB/4D5B
  • PDB/4D5D
  • PDB/4V3G
  • PDB/4V3H