3 DIFFERENT APPROACHES FOR CALCULATING THE 3-DIMENSIONAL STRUCTURE OFMICROTUBULES DECORATED WITH KINESIN MOTOR DOMAINS

We have used three different electron microscopy approaches to calcula te three-dimensional maps of tubulin assemblies decorated with the mot or domain of kinesin. The approaches used were: (1) Tilt series recons truction of negatively stained tubulin sheets. (2) Back-projection rec onstruction of microtubules in ice. (3) Helical reconstruction of micr otubules in ice. The calculated maps show the overall configuration of the protofilaments and the interactions between the motor and the pro tofilaments at a resolution of 2-4 nm. The three methods revealed a si milar binding configuration of the kinesin motor domain to the protofi lament. We also found that seams can be present in potentially helical microtubules, limiting the use of helical reconstruction algorithms. Advantages and disadvantages of each of the three approaches are discu ssed. (C) 1997 Academic Press.