UNIT 17.2 Electron Microscopy and Image Processing: An Essential Tool for Structural Analysis of Macromolecules

Abstract

Macromolecular electron microscopy (EM) deals with macromolecular complexes and their placement within the cell—linking the molecular and cellular worlds as a bridge between atomic-resolution X-ray crystallographic or NMR studies and lower resolution light microscopy. The amount of specimen required is typically 10² to 10³ times less than for X-ray crystallography or NMR. Electron micrographs of frozen-hydrated specimens portray native structures. Computer averaging yields enhanced images with reduced noise. Three-dimensional reconstructions may be computed from multiple views. Under favorable circumstances, resolutions of 7 to 10 Å are achieved. Fitting atomic-resolution coordinates of components into three-dimensional density maps gives pseudo-atomic models of a complex's structure and interactions. Time-resolved experiments describe conformational changes. Electron tomography allows reconstruction of pleiomorphic complexes and sub-cellular structures. Electron crystallography has produced near-atomic resolution models of two-dimensional arrays, notably of membrane proteins.