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Gram-negative trimeric porins have specific LPS binding sites that are essential for porin biogenesis

DOI: 10.1073/pnas.1602382113 DOI Help

Authors: Wanatchaporn Arunmanee (Institute for Cell and Molecular Biosciences, Newcastle University) , Monisha Pathania (Institute for Cell and Molecular Biosciences, Newcastle University) , Alexandra S. Solovyova (Newcastle University) , Anton P. Le Brun (National Deuteration Facility, Bragg Institute, Australian Nuclear Science and Technology Organisation) , Helen Ridley (Institute for Cell and Molecular Biosciences, Newcastle University) , Arnaud Basle (Institute for Cell and Molecular Biosciences, Newcastle University) , Bert Van Den Berg (Newcastle University) , Jeremy H. Lakey (aInstitute for Cell and Molecular Biosciences, Newcastle University)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Proceedings Of The National Academy Of Sciences , VOL 113 , PAGES E5034 - E5043

State: Published (Approved)
Published: August 2016
Diamond Proposal Number(s): 9948

Open Access Open Access

Abstract: The outer membrane (OM) of gram-negative bacteria is an unusual asymmetric bilayer with an external monolayer of lipopolysaccharide (LPS) and an inner layer of phospholipids. The LPS layer is rigid and stabilized by divalent cation cross-links between phosphate groups on the core oligosaccharide regions. This means that the OM is robust and highly impermeable to toxins and antibiotics. During their biogenesis, OM proteins (OMPs), which function as transporters and receptors, must integrate into this ordered monolayer while preserving its impermeability. Here we reveal the specific interactions between the trimeric porins of Enterobacteriaceae and LPS. Isolated porins form complexes with variable numbers of LPS molecules, which are stabilized by calcium ions. In earlier studies, two high-affinity sites were predicted to contain groups of positively charged side chains. Mutation of these residues led to the loss of LPS binding and, in one site, also prevented trimerization of the porin, explaining the previously observed effect of LPS mutants on porin folding. The high-resolution X-ray crystal structure of a trimeric porin–LPS complex not only helps to explain the mutagenesis results but also reveals more complex, subtle porin–LPS interactions and a bridging calcium ion.

Journal Keywords: gram-negative bacteria; lipopolysaccharide; X-ray crystal structure; porin; outer membrane

Subject Areas: Biology and Bio-materials

Instruments: I02-Macromolecular Crystallography

Other Facilities: Institut Laue-Langevin; ESRF