Three‐dimensional reconstruction from a single‐exposure, random conical tilt series applied to the 50S ribosomal subunit of Escherichia coli

Abstract

SUMMARY: We present a new reconstruction method that takes advantage of the fact that many biological macromolecular assemblies show a preferred orientation with respect to the plane of the specimen grid in the electron microscopic preparation. From one micrograph taken of such a specimen tilted by a large angle, a conical tilt series with random azimuthal angles can be extracted and used for a three‐dimensional reconstruction. Our technique allows the determination of the molecular structure under low‐dose conditions, which are not achievable with reconstruction methods that use conventional tilt series. The reconstruction method combines a number of existing image processing techniques with a newly developed weighted back‐projection algorithm designed for three‐dimensional reconstruction from projections taken with arbitrary projecting directions. The method is described as it was applied to the three‐dimensional reconstruction of the structure of the 50S ribosomal subunit of Escherichia coli (E. coli).