Visualizing large macromolecular assemblies in vivo with electron cryotomography

Abstract

Electron cryotomog. (ECT) has made it possible to visualize large macromol. assemblies inside intact cells in a near-native, "frozen-hydrated" state in 3-D to 2-5 nm resoln. Increasingly, at. models of individual proteins and smaller complexes obtained by X-ray crystallog., NMR spectroscopy, or other methods can be fit into cryotomograms to reveal how the various pieces work together inside cells. I will illustrate the potential of ECT by presenting our recent work on two macromol. machines involved in bacterial motility: chemoreceptor arrays and the flagellar motor. Flagellar motors share a conserved core, but exhibit remarkably diverse adaptations in different species. Cryotomograms of knockout strains are now revealing the locations of various protein components including FliI, FliH, and FlhA. In contrast, chemoreceptor arrays are universally conserved, hexagonallypacked trimers-of-receptor-dimers, linked together by rings of CheA and CheW in a way that suggests new mechanistic models for their exquisite sensitivity and adaptability.