1] Studying ribosome structure by electron microscopy and computer-image processing

Abstract

This chapter focuses on the study of ribosome structure by electron microscopy and computer-image processing. In the method of single-particle 3-D reconstruction the macromolecule is reconstructed in three dimensions from a large number of projections. In the variant of this method most viable for structure research, these projections come from different particles lying in, or brought into, different orientations providing a wide range of viewing directions. In principle, electron crystallographic techniques are capable of providing structural data on ribosomes. However, a resolution comparable to that obtained by single-particle reconstruction has not been achieved, mainly because large, well-ordered crystals of ribosomes or ribosomal subunits have been difficult to grow. The statistical significance of structural features in an averaged image can be assessed by applying standard statistical tests to the individual image elements. The theoretical resolution of a 3-D reconstruction, which depends on the number of projections, may be calculated for equal angular increments in both the single-axis and the conical-tilting schemes. Both formulas are derived by using Shannon's sampling theorem, which states in essence that two measurements must be available for each resolution elements.