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![[Figure 1]](id5005fig1mag.jpg)
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Figure 1
GMPCPP-stabilized in vitro-polymerized MTs visualized by cryo-electron tomography. (a) Top, 0° tilt image (motion-corrected movie sum) from a tilt series, showing a typical field of MTs that includes views of the MT lattice (red arrowhead, including the moiré pattern) and MT ends (pink arrow), together with 10 nm nanogold fiducial markers (black arrows). Bottom, 8 nm section through the three-dimensional tomographic reconstruction of the same sample, showing the appearance of sections through the MT wall (red arrowhead) in which individual PFs are visible, the MT lumen (orange arrowhead) and MT ends (pink arrow). Inset: Fourier transform of an extracted MT showing a layer line corresponding to the 4 nm spacing of the tubulin monomers within the MT lattice, indicative of the level of structural detail in these data. (b) An exemplar MT image (0° tilt image; left) and Fourier filtered image (right) showing the moiré pattern characteristic of a 14PF MT (black arrows). The arrowhead shapes within the moiré pattern (red arrowheads) indicate the MT polarity, with the MT plus end towards the top in this case. (c) Rotational averaging analysis of 8 nm thick transverse sections from the tomographic reconstruction supports the moiré-pattern-based assignment of 14PF architecture for this MT, yielding clear PF projections in the 14-fold average, while 13-fold or 15-fold averaging of the same data blurs out structural information from the characteristic PF projection. (d) Transverse section (top) and segmented and surface-rendered view (bottom) through the same three-dimensional tomogram, showing the distribution of the MTs (red) in the vitreous ice layer (indicated by the shaded area) and the tendency of the background protein (tubulin, yellow) to lie at the air–water interface of the sample. This view also shows the distorting effect of the missing wedge on the MT structure, illustrating the difficulty of directly counting PFs in such data. |
![[Figure 1]](id5005fig1.jpg)
 | STRUCTURAL BIOLOGY |
ISSN: 2059-7983
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