HI 6326 Session 7

Overview:

  1. Lecture 1: 3D Electron Microscopy History (PDF file)
  2. Video Lecture: Michael Moody, Helical Reconstructions (640x480 RealVideo, 5 fps, 620MB)
  3. Homework 2

Resources and Assignments:

1. If you are interested in cryo-EM Specimen Preparation (not discussed in Lecture 1):
     Note: References 6 and 10 in the Butcher review are in the Biomachina lab library

2. Here are the PDF's of the historic papers and reviews mentioned in Lecture 1:
 3. The handout of the Video Lecture can be downloaded as a PDF file here.

4. Homework 2 (due at last session):

Background:

Actin filaments are dynamic polymers whose ATP-driven assembly in the cell cytoplasm drives shape changes, cell locomotion and chemotactic migration. Actin filaments also participate in muscle contraction. The structure of the filament is not known at atomic resolution, but several models were produced in the laboratory of Ken Holmes (Max-Planck Institute for Medical Research, Heidelberg, Germany) by refinement against X-ray fiber diffraction data:


Fig. 1 (click to enlarge): A single actin monomer with inter-actin contact surfaces is shown on the right, the entire actin filament on the left. The figure is copyrighted by W.W.

Assignment:

Read the hands-on practical guide for helical indexing by David DeRosier.  Using the helical selection rule, draw a "n-l" plot for actin filaments in both 13/6 symmetry (l= -6n+13m) and 36/17 (l= -17n+36m) symmetry (see section 12., page 10).  Actin can be found in both symmetries (and other ones!) because there is some torsional flexibility in the filaments that can accommodate a variable twist. Now, draw an "n-Z" plot, assuming that one actin subunit has a rise of 27.5 Angstrom. Calculate the Z value (length of helical repeat) l=1 corresponds to, for 13/6 and 36/17. Then normalize the "l" axis in both cases to get the Z axis. See Figure 5, page 15, in the handout as an example of an n-Z plot. Note that the twist angle per monomer is obviously -170 degrees for 36/17 symmetry. Since this angle is close to -180 degrees, actin seems to have the appearance of a right-handed two-start helix instead of a left-handed one-start helix. Calculate how this twist angle changes for 13/6 symmetry. You should verify that despite the large difference in indexing and n-l plots we really have very similar filaments here, so their diffraction patterns should also be very similar. Can you find this similarity in the n-Z plots?