Fig. 4: Emulation of actin filament by D-loop grafting.

a Schematic representations of PuuE(D-loop)-M. The position of the D-loop graft (red) is indicated by protein sequence (top) and the AF2-predicted structure (bottom). Symmetry of protein is represented with lines and square markers. b nsTEM images of tubes with a helical conformation composed of PuuE(D-loop)-M and PuuE-p. The helical pattern of two (centre) or three (right) intertwined tubes is shown in the high-magnification image. c nsTEM images showing the reversibility of the tube structure with helical conformations by temperature change. Temp. indicates temperature. d Representative cryo-EM images (top) and 2D class-averaged images (bottom) of helical tube structures. e Representative cryo-EM image (top) and 2D class-averaged image of tube structure with C₃ symmetry. Tube structures with other symmetries found in this study are shown in Supplementary Fig. 10. f 3D reconstructed model of the tube structure with C₃ symmetry. For visibility, only the PuuE structure (PDB ID: 3CL6) is overlayed on the 3D reconstructed model. g Fitting of AF2-predicted model of PuuE-p into the 3D reconstructed model. The fitting results suggest that PuuE-p is unlikely to fit in the units located inside the tube structure; it is better accommodated by the units on the outside. The map density was normalised and visualised at a threshold of sigma (σ) value = 3.3 above average. Structure fitting was performed to count the atoms of the PuuE-p model outside the reconstructed 3D map of the C₃ tube structure at this threshold. Based on this prediction, the units in f are colour-coded as described in Fig. 1c. The Met-6xHis-TEVcs region of the PuuE-p model is not shown to improve visibility. Scale bars, 1 μm (white), 100 nm (black), 10 nm (grey).