Journal: bioRxiv

Article Title: Crystal structure of the membrane anchoring domain of mycobacterial Wag31: a dimer-of-dimer suggests how a Wag31 filament might self-assemble

doi: 10.1101/869594

Figure Lengend Snippet: (a) Two nearly-orthogonal views of N-Wag31 (2-60 residues) dimer as blue ribbons are shown. The calculated local coiled coil axes are shown as red jointed lines. (b) CD profile (molar residue ellipticity versus wavelength) of N-Wag31. (c) Superimposed structures of N-Wag31 (blue) and N-bsDivIVA (coral) are shown. (d) The intertwined loop region in N-Wag31 (blue) containing the P 16 P 17 I 18 sequence (ball and stick, with carbon atoms in light blue, oxygen in red and nitrogen in blue), and equivalent region in N-bsDivIVA (coral) containing the stacked F17 residues (ball and stick, with carbon atoms in coral, oxygen in red and nitrogen in blue), are shown. (e-g) Three views of the tetrameric N-Wag31 (grey) with electrostatic potential mapped into the water-accessible surface (water radius 1.4 Å) are shown. Electrostatic potential was computed using APBS with AMBER charges and suggested parameters . Figures were prepared using Pymol (The PyMOL Molecular Graphics System, Schrodinger, LLC.) and CCP4MG from the CCP4 suite .

Article Snippet: The N-terminal domain of Wag31 (2-60 aa with an N-terminal tag containing poly-histidine residues, synthesized by Genescript) construct in pET15b vector was expressed in Escherichia coli BL21 (DE3) cells.

Techniques: Residue, Sequencing

Journal: bioRxiv

Article Title: Crystal structure of the membrane anchoring domain of mycobacterial Wag31: a dimer-of-dimer suggests how a Wag31 filament might self-assemble

doi: 10.1101/869594

Figure Lengend Snippet: Structure of N-Wag31. The monoclinic unit cell is shown as a box. N-Wag31 dimer within the asymmetric unit, including the poly-histidine tag regions (see methods section), is shown as a cartoon.

Article Snippet: The N-terminal domain of Wag31 (2-60 aa with an N-terminal tag containing poly-histidine residues, synthesized by Genescript) construct in pET15b vector was expressed in Escherichia coli BL21 (DE3) cells.

Techniques:

Journal: bioRxiv

Article Title: Crystal structure of the membrane anchoring domain of mycobacterial Wag31: a dimer-of-dimer suggests how a Wag31 filament might self-assemble

doi: 10.1101/869594

Figure Lengend Snippet: (a-b) Two views of the dimer-of dimer of N-Wag31 related by crystallographic 2-fold symmetry are shown as ribbons (cyan and grey). The interface-forming H2 helices are marked. A PGE molecule bound to N-Wag31 is shown as sticks in (b). (c) Tetrameric N-Wag31 is shown as ribbon (cyan) within the molecular surface (grey) of the tetramer. Side-chains of the interface forming residues R41, E27, L34, A30 and E38 are shown in ball-and-stick (carbon in magenta, oxygen in red and nitrogen in blue). (d) Multiple sequence alignment of the N-term region of tbWag31 (numbering for tbWag31 in red) and homologs, with the degree of conservation, is shown (prepared in Jalview, ). (e) SAXS elution profile (average intensity versus frame number obtained from CHROMIXS, ) of N-Wag31. Guinier plot (lnI(q) versus q 2 , I is intensity and q is momentum transfer in nm −1 ) for peak II is shown in the inset. (f) Scattering profile of peak II data (I versus q in Å −1 ). Computed scattering profiles of dimeric and tetrameric N-Wag31 (calculated using CRYSOL with suggested parameters, ) are shown fitted to the experimental data.

Article Snippet: The N-terminal domain of Wag31 (2-60 aa with an N-terminal tag containing poly-histidine residues, synthesized by Genescript) construct in pET15b vector was expressed in Escherichia coli BL21 (DE3) cells.

Techniques: Sequencing

Journal: bioRxiv

Article Title: Crystal structure of the membrane anchoring domain of mycobacterial Wag31: a dimer-of-dimer suggests how a Wag31 filament might self-assemble

doi: 10.1101/869594

Figure Lengend Snippet: (a-b) Two theoretically alternative possible modes of self-assembling of Wag31, using (a) pure translational repeat, and (b) two-fold symmetry related repeat, are shown. (c) Ribbon diagrams of the tetramers of N-terminal N-Wag31 (blue) and C-terminal domain of bsDivIVA (red) are shown, joined by linker region as dashed lines, as a theoretical filament. The linker region is ∼20 residue long in bsDivIVA, and ∼100 residue long in tbWag31. (d) A theoretically possible mode of 3-way branching in tbWag31 filament. (e-f) Two views of a theoretical hexamer composed of three dimers of N-Wag31 (ribbons in green, blue and magenta). The two-fold axes of rotation are shown as red rods.

Article Snippet: The N-terminal domain of Wag31 (2-60 aa with an N-terminal tag containing poly-histidine residues, synthesized by Genescript) construct in pET15b vector was expressed in Escherichia coli BL21 (DE3) cells.

Techniques: Residue