Chapter H2 - Three-dimensional reconstruction from two-dimensional images

Summary

EM in biology

Structural biology with EM

Electron microscopes can provide vivid details of biological macromolecules at the nanometre scale.

Recall from Chapter H1, an electron image is a two-dimensional projection of the object along the electron beam. A three-dimensional reconstruction of the object can be obtained by combining data measured at different angular projections. In single-particle reconstruction, a population of molecules is imaged. If these are identical (monodisperse) and present in sufficient quantities, the data from differently oriented molecules can be combined to produce an accurate three-dimensional reconstruction of the structure. The presence of symmetry is taken advantage of in helical reconstruction and two-dimensional crystallography. In tomography, the same object is imaged at different orientations. The different images are then combined together to recreate its structure.

Examples of electron cryo-microscopy reconstructions

In EM, the image is the result of the interaction of the incident electrons with the electrostatic distribution due to the atomic structure of the sample. The reconstruction of the structure results from the analysis of many images. Three-dimensional reconstructions at different resolutions depict the object at different levels of detail (Figure H2.1).

At atomic resolution, it is a good approximation to understand this structural density as being equivalent to the electron density obtained by X-ray diffraction, but with a larger contribution from hydrogen atoms in the electron diffraction case. It should furthermore be possible to distinguish the charge state of amino acids.