This is a request for a 200 kV electron microscope (EM) with a field-emission gun, a liquid-nitrogen-cooled stage, and a 4K CCD detector. Structural studies of large macromolecular complexes have become an important and expanding research focus in laboratories at Harvard Medical School and its affiliated institutions, and molecular electron microscopy has become a central tool of many research groups. This rapidly growing interest has generated an evident need for the instrument requested. It is now possible to get sub-nanometer resolution density maps from single-particle analysis of images from cryo-stabilized samples. Secondary-structural features are clearly detectable at those resolutions, providing an accurate link between high-resolution structures of protein components (e.g., from x-ray crystallography) and EM image reconstructions of much larger assemblies of those components. Research projects from seven groups (Dormitzer, Ellenberger, Harrison, Hogle, Kirchhausen, Rapoport, Walz) are outlined. The goals include: structural basis of viral entry into cells, DNA remodeling enzymes, assembly and uncoating of clathrin coated vesicles, protein translocation, structure of intercellular membrane junctions, structure of iron-transporting proteins, structure of the spliceosome. Approaches include both single-particle analysis and 2D electron crystallography. ? ?
| Norville, Julie E; Kelly, Deborah F; Knight, Thomas F et al. (2011) Fast and easy protocol for the purification of recombinant S-layer protein for synthetic biology applications. Biotechnol J 6:807-11 |
