CRYO TRANSMISSION ELECTRON MICROSCOPE

The users assembled in this application have a common interest in using cryo transmission electron microscopy (cryoTEM) of vitreous ice- embedded samples as a quantitative tool to investigate the assembly, regulation and function of biologically important macromolecular complexes. Despite continuous progress in x-ray crystallographic and NMR methods, it is still difficult using these techniques alone to obtain atomic level structural information about the many complex macromolecular assemblies that govern fundamental cell biological processes. It is now clear that a powerful approach is to combine structural information obtained at different levels. The structural information obtained from cryoTEM and computer based 3-D reconstruction spans the gap between our ability to obtain structural information about large macromolecular assemblies using conventional light and electron microscopy, and high-resolution data obtained from NMR or X-ray crystallography. This data can be combined to build up a detailed picture of the overall architecture of the complexes and the interactions between the components. This grant is based on projects in which such a complimentary approach is used to analyze complex biological structure. The major users in this application will use the requested cryo-TEM to probe file mechanisms of actin binding protein interactions (Almo, Condeelis), bacterial flagellar motor switching (Khan) and virus entry/exit (Kielian) at the molecular level. At the level of ultrastructure, the developmental program of C. elegans will be elucidated by examination of its fine morphology as revealed in cryo-sections (Hall).