Lab Overview
Macromolecular assemblies may be composed of from two to perhaps scores of
proteins and are the functional units – the molecular machines - of the
cell. We use cryo-electron microscopy and image analysis to study the structure
and mechanism of action of several of these machines. The three-dimensional
maps we calculate from electron images of the machines are combined with data
from other sources to provide insight into their operation. During the past
year, we have examined the mechanisms of microtubule stabilization and destabilization
by several proteins, and continued our work on backwards-moving kinesins and
myosins, VCP/p97, bacterial toxins and membrane proteins.
Highlight 2004
Mechanism of minus-end directed movement along microtubules by kinesin 14s.
Craig Yoshioka, Nick Endres, Ron Vale, Ron Milligan.
A unique class of kinesin motor proteins (Kinesin 14) powers movement towards
the microtubule minus end. The mechanism of motility for these minus-end kinesins
is poorly understood in comparison to the plus-end-directed kinesins. We have
used cryo-electron microscopy to visualize a 75°, minus-end-directed rotation
of the mechanical element of the Kinesin 14 motor NCD upon binding of ATP. Extending
or shortening the mechanical element increases or decreases movement velocity
respectively without affecting ATPase activity. These results show that, like
other kinesins, NCD’s force-producing conformational change occurs upon
ATP binding but involves a lever-arm-like swing similar to that described for
myosins.
2004 publications
Carragher, B., D. Fellmann, F. Guerra, R.A. Milligan, F. Mouche, J. Pulokas,
B. Sheehan, J. Quispe, C. Suloway, Y. Zhu and C.S. Potter. 2004. Rapid routine
structure determination of macromolecular assemblies using electron microscopy:
current progress and further challenges. J. Synchrotron Radiation, 11: 83-85
Roger, B., J. Al-Bassam, L. Dehmelt, R. Milligan, and S. Halpain. 2004. MAP2c,
but not tau, binds and bundles f-actin via its microtubule binding domain,
Current Biol., 14:363-71.
Batters, C., C.P. Arthur, A. Lin, J. Porter, M.A. Geeves, R.A. Milligan,
J.E. Molloy and L.M. Coluccio. 2004. Myo1c is designed for the adaptation
response
in the inner ear. EMBO J., 23:1433-40.
