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

Abstract 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. Supplementary information Supplementary data are available at Bioinformatics online.

: Pre-and post-kink helix axes are plotted on an overlay of the 2ahy structure (one chain is depicted in red with circular capped axes) and 3e83 (one chain is depicted in green with diamond capped axes). The superposition was performed using the PyMol "super" command. The pore axis is shown as a dotted line in the left panel and a crossed circle in the right panel (90 degree view).
Supplementary Figure S1b: The internal surface dimensions of the NaK channel pore as calculated using HOLE (Smart et al, 1996), and depicted as a wire mesh inside the closed (PDBID 2ahy, left) and open (PDB: 3e83, right) structures, with the chains coloured as in Figure S1a.

Example 2: Comparison of open and closed channel structures
The prokaryotic ligand gated ion channel (GLIC) is a homopentamer which forms a proton-gated cation channel. GLIC is of particular interest because of its homology to eukaryotic acetylcholine receptors; like acetylcholine receptors GLIC is a target of general anesthetics. Co-crystallization of GLIC with a general anesthetic produced a closed form of the channel (PDBID 4zzb) which can be used for direct comparison with the open channel (PDBID 4zzc) to investigate the structural changes involved in gating (Sauguet et al, 2016). AnglerFish was used to calculate (Table S2)   Supplementary Figure S2: Helical axes plotted on the overlaid 4zzb (in red with diamond capped axes) and 4zzc (in green with circle capped axes) structures; the helices are numbered as in Table S2. Superposition was performed using the Pymol "super" command. The pore axis is shown as a dotted line and circled cross. The numbers correspond to the order of the helices from N-to C-termini.

Example 3: Analysis of the asymmetric opening of the NavMs pore:
The voltage-gated sodium channel from Magnetococcus Marinus (NavMs) is a homotetramer and is used as a structural model for eukaryotic Navs. The first open structure sodium channel structure (McCusker et al, 2012) solved that was of the NavMs pore. This structure showed the pore to be asymmetric, with the gate partially open, as a consequence of a different psi angle in one (chain A) of the S6 helices monomers. A later structure of the NavMs pore (Bagneris et al, 2013) depicted the symmetric conformation of a fully open pore.
In order to demonstrate the asymmetry of the partially-open pore (PDBID 4f4l), tilt and swing values were calculated for each chain. Because chain A corresponded to the altered conformation, chains B, C and D were used to define the pore axis; had all chains been used, they would have skewed the orientation of the axis. Tilt and swing values for chain A were 141.0° and 27.2°, respectively, which fall outside the s.   Figure S3: Partially (red) and full (green) open NavMs structures overlaid, with the chains letters as in Table S3. The helical axis for chain A is shown as the solid black line, and highlights that the A chains in both structures are the same, but the other chain are not. The superposition was performed using the Pymol "super" command. The pore axis is shown as a dotted line in the left panel and a circled cross in the right panel, which was produced by a 90 degree rotation of the figure in the left panel.