The bacterial flagellar filament is a hydrodynamic propeller that converts the motion of the rotary motor into linear thrust. The canonic flagellin monomer (Salmonella typhymurium SJW1655) is comprised, from the outside in, of four domains D0-D3. It assembles into a helical polymer with major helical lines of orders n= 1, -5,+6 and ±11.
Non-helical perturbations are known to occur in S. typhimurium SJW23 and SJW117, which are straight, right- handed and left-handed filaments, respectively. These perturbations are a product of pairwise symmetry reduction along the left-handed, five start (-5) helical lines. The resulting helical lines can be indexed only as fractional Bessel orders (-2.5, -13.5, 8.5, 3.5 etc), which indicate disruption of the continuity of the helical lines (and a consequent seam).
Previously we attempted to determine the molecular nature and origin of the non-helical perturbation: (a) by perturbing the plain structure of a model so that a power spectrum similar to that of SJW117/23 is obtained. This is achieved by a ±10° rotation of alternating globular subunit rows along the 5-start lines. (b) by ‘reverse engineering’ the SJW23 flagellin gene (fliC23) and restoring the filament’s ‘plain’ helical symmetry (as indicated by changes in the power spectrum--disappearance of the typical layer-line clusters (n= -2.5, -13.5, 8.5)). We concluded that the non-helical perturbation is a product of unique interactions in the inner part of the D3 density shell (contrary to interactions at the outer tip of D3 in the case of helical perturbations).
However, without a full three dimensional reconstruction we cannot specify the spatial and molecular interactions leading to the perturbation. The non integer Bessel orders introduce helical symmetry breaking in the form of a seam which prevents the straightforward implementation of symmetry-based Fourier-Bessel and iterative real-space three-dimensional image reconstruction techniques. Methods exist to reconstruct microtubules with seams; however, these are thin walled, relatively straight tubes assembled from relatively simple globular monomers.
We refined real-space iterative helical reconstruction methods suitable for handling the unique properties of non-helically perturbed bacterial flagellar filaments, which are complex, dense, helical polymers, constructed from multi-domain monomers resulting in large fluctuations in radial mass distribution which allow for complex perturbations. We reconstruct the filament without using a reference (similar to IHRSR) and bring into account the relation between the seam‘s angular position and the tube’s radius.
Our findings suggest that domain D2 is the anchor of the non-helical perturbation. It allows a sharp tilt of D2, D3 and the reduction in symmetry (dimerization): the connectivities between D2 with the inner part of D3 gives rise to helical lines of order n= -2.5 and -13.5. The tilted orientation of D2, D3 can be explained by the hydrophilic nature of a modified α-helix at the interface of D2 with D1. Another modification of a α-helix, at the interface of D2 with the neighboring D1 gives rise to the helical lines of orders n= -2.5. D2/D2 interactions give rise to n=8.5 lines. These interactions allow for the dimerization of monomers.
