Lipid binding attenuates channel closure of the outer membrane protein OmpF.

Strong interactions between lipids and proteins occur primarily through association of charged headgroups and amino acid side chains, rendering the protonation status of both partners important. Here we use native mass spectrometry to explore lipid binding as a function of charge of the outer membrane porin F (OmpF). We find that binding of anionic phosphatidylglycerol (POPG) or zwitterionic phosphatidylcholine (POPC) to OmpF is sensitive to electrospray polarity while the effects of charge are less pronounced for other proteins in outer or mitochondrial membranes: the ferripyoverdine receptor (FpvA) or the voltage-dependent anion channel (VDAC). Only marginal charge-induced differences were observed for inner membrane proteins: the ammonia channel (AmtB) or the mechanosensitive channel. To understand these different sensitivities, we performed an extensive bioinformatics analysis of membrane protein structures and found that OmpF, and to a lesser extent FpvA and VDAC, have atypically high local densities of basic and acidic residues in their lipid headgroup-binding regions. Coarse-grained molecular dynamics simulations, in mixed lipid bilayers, further implicate changes in charge by demonstrating preferential binding of anionic POPG over zwitterionic POPC to protonated OmpF, an effect not observed to the same extent for AmtB. Moreover, electrophysiology and mass-spectrometry-based ligand-binding experiments, at low pH, show that POPG can maintain OmpF channels in open conformations for extended time periods. Since the outer membrane is composed almost entirely of anionic lipopolysaccharide, with similar headgroup properties to POPG, such anionic lipid binding could prevent closure of OmpF channels, thereby increasing access of antibiotics that use porin-mediated pathways.

AnglerFish: a webserver for defining the geometry of α-helices in membrane proteins.

Summary: Integral membrane proteins that form helical pores and bundles constitute major drug targets, and many of their structures have been defined by crystallography and cryo-electron microscopy. The gating of channels and ligand binding of transporters generally involve changes in orientation of one or more the constituent helices in the structures. At present there is no standard easily accessible means for defining the orientation of a helix in a membrane protein structure. AnglerFish is a web-based tool for parameterising the angles of transmembrane helices based on PDB coordinates, with the helical orientations defined by the angles 'tilt' and 'swing'. AnglerFish is particularly useful for defining changes in structure between different states, including both symmetric and asymmetric transitions, and can be used to quantitate differences between related structures or different subunits within the same structure. Availability and Implementation: AnglerFish is freely available at http://anglerfish.cryst.bbk.ac.uk . The website is implemented in Perl-cgi and Apache and operation in all major browsers is supported. The source code is available at GitHub. Contact: b.wallace@mail.cryst.bbk.ac.uk. Supplementary information: Supplementary data are available at Bioinformatics online.

Dimerization of complement factor H-related proteins modulates complement activation in vivo.

The complement system is a key component regulation influences susceptibility to age-related macular degeneration, meningitis, and kidney disease. Variation includes genomic rearrangements within the complement factor H-related (CFHR) locus. Elucidating the mechanism underlying these associations has been hindered by the lack of understanding of the biological role of CFHR proteins. Here we present unique structural data demonstrating that three of the CFHR proteins contain a shared dimerization motif and that this hitherto unrecognized structural property enables formation of both homodimers and heterodimers. Dimerization confers avidity for tissue-bound complement fragments and enables these proteins to efficiently compete with the physiological complement inhibitor, complement factor H (CFH), for ligand binding. Our data demonstrate that these CFHR proteins function as competitive antagonists of CFH to modulate complement activation in vivo and explain why variation in the CFHRs predisposes to disease.