Variations of the three-dimensional structure of the Escherichia coli ribosome in the range of overlap views. An application of the methods of multicone and local single-cone three-dimensional reconstruction

Abstract

Electron microscopic techniques are among the most important tools for obtaining structural information of biological specimens. However, the three-dimensional (3D) structural analysis of asymmetrical specimens that do not form crystalline sheets has traditionally presented serious methodological obstacles to its accomplishment. One of the fundamental questions to be addressed in this type of structural study is in what way, and to what degree, does the 3D structural conformation depend on the orientation of the specimen with respect to the electron microscopic support films. As a step in studying this problem, we have analyzed the variations of the 3D structure of the Escherichia coli 70S monosome by performing four different 3D reconstructions of the 70S monosome from subsets of images in the so-called overlap range of views. These subsets were selected according to a multivariate statistical analysis performed on the total population of overlap-range specimen images. A certain amount of structural variability exists among the 3D reconstructions, although many of the main morphological characteristics, as the relative orientation between the ribosomal subunits, remain unchanged. We have also generalized the random conical reconstruction technique (Radermacher, M., T. Wagenknecht, A. Verschoor, and J. Frank. 1987. J. Microsc. 146: 113-136) to include those cases where the specimen exhibits a rocking behavior with respect to the support. The resulting Multicone Reconstruction Technique has been applied to computer-generated images as well as the E. coli 70S monosome images from part of the overlap range of views.