P. N. T. Unwin

Three-dimensional model of purple membrane obtained by electron microscopy

A 7-Å resolution map of the purple membrane has been obtained by electron microscopy of tilted, unstained specimens. The protein in the membrane contains seven, closely packed, α-helical segments which extend roughly perpendicular to the plane of the membrane for most of its width. Lipid bilayer regions fill the spaces between the protein molecules.

Molecular structure determination by electron microscopy of unstained crystalline specimens.

Abstract The projected structures of two unstained periodic biological specimens, the purple membrane and catalase, have been determined by electron microscopy to resolutions of 7 A and 9 A, respectively. Glucose was used to facilitate their in vacuo preservation and extremely low electron doses were applied to avoid their destruction. The information on which the projections are based was extracted from defocussed bright-field micrographs and electron diffraction patterns. Fourier analysis of the micrograph data provided the phases of the Fourier components of the structures; measurement of the electron diffraction patterns provided the amplitudes. Large regions of the micrographs (3000 to 10,000 unit cells) were required for each analysis because of the inherently low image contrast ( Our methods appear to be limited in resolution only by the performance of the microscope at the unusually low magnifications which were necessary. Resolutions close to 3 A should ultimately be possible.

Electron microscopy of the stacked disk aggregate of tobacco mosaic virus protein: II. The influence of electron irradiation on the stain distribution

Abstract Optical and computer analysis of bright field electron micrographs of the same negatively stained stacked disk specimens subjected to different electron doses shows that the stain migrates and redistributes itself over the protein surfaces during irradiation. The analysis suggests that the main factors responsible for this migration are the contraction of the stain inevitably brought about by the irradiation, and associated surface energy effects. The morphology of the protein has a strong influence on the path of the migration. A major consequence of these changes in stain distribution is that conventional bright field micrographs taken under normal dosage conditions tend to give a somewhat misleading representation of the specimens original structure. Migration of the stain under the electron beam is likely to be a fairly general phenomenon and it therefore seems important to take bright field micrographs of negatively stained specimens using the smallest possible electron doses.

Beef liver catalase structure: Interpretation of electron micrographs

It is shown that the catalase platelets frequently examined by electron microscopists have the space group symmetry P212121 and contain four molecules in the orthorhombic unit cell. The cell dimensions, determined from X-ray diffraction patterns, are: a=69 A, b=173·5 A, c=206 A. Comparison of images of the platelets, prepared in different ways, indicates that the normal negative staining pattern formed by them gives only a very rough, low resolution representation of their native structure.

The origin of the grain boundary precipitate free zone

Abstract The origin of the grain boundary precipitate free zone is discussed in relation to Al-Zn-Mg and Al-Cu alloys using the occurrence of homogeneously distributed precipitate as a semi-quantitative indication of the excess vacancy concentration and its variation in the quenched alloys. It is shown that all boundaries except those with a misorientation 2° act as ideal sinks for vacancies. In order to explain the occurrence of the precipitate free zone and its behaviour under different heat treatment conditions it is necessary to consider: (a) the presence of a vacancy concentration profile in the quenched alloy resulting from loss of vacancies to the grain boundary and, on occasions, to intragranular sinks such as dislocation loops; (b) the presence of a solute concentration profile in cases where grain boundary precipitation has occurred as a result of a slow quench or a quench interruption treatment.

The nucleation and initial stages of growth of grain boundary precipitates in Al-Zn-Mg and Al-Mg alloys

Abstract The early stages of grain boundary precipitation in two aluminium alloys, Al-6 %Zu-3 %Mg and Al-7 % Mg, have been examined by transmission electron microscopy, and are discussed in relation to the heat treatment procedure and the properties of individual boundaries. The precipitates may be nucleated in two stages; one associated with quenching, and the other with the ageing treatment. The quench rate, the solution treatment temperature and the ageing treatment are found to be the important parameters influencing precipitation behaviour. It has also been found that the nucleation and growth of boundary precipitates in these alloys is strongly affected by the type of boundary and the results have been interpreted using the coincidence site model of grain boundaries.

Two forms of isolated gap junctions

Abstract Gap junctions, containing regular hexagonal arrays of connexons, have been isolated from rat liver. The projected structures of these gap junctions have been studied to a resolution of 18 A by electron microscopy of negatively stained samples. Two closely related forms of junction were produced that have different structures for the connexon, but the same hexagonal lattice constant. In one form the connexon is seen as a weakly contrasted annulus, which is broadest at the locations where other connexons come closest; in the other, the connexon is seen as a strongly contrasted annulus, which is broadest midway between the locations where other connexons come closest. The forms appear to reflect two configurations of the connexon subunits.

Electron microscopy of the stacked disk aggregate of tobacco mosaic virus protein: I. Three-dimensional image reconstruction

Abstract The three-dimensional structure of the stacked disk aggregate of tobacco mosaic virus protein has been determined from “phase plate” electron micrographs to an effective resolution of about 12 A. It is a long rod comprised of paired rings of protein (disks), the subunits of which have different conformations according to which ring they belong. The two subunit conformations are such that the rings come close together within a disk near the outer surface of the particle, but between disks on the inside. This property, interpreted on the basis of a polar packing of the subunits, was established from an earlier, lower resolution, study by Finch & Klug (1971). The present study shows, in addition, that the pairing is contributed mainly by axial distortions of the subunits in one of the rings, the axial distortions of the subunits in the other being largely replaced at lower radii by a tilt or twist and, at higher radii, by a slew. The subunits in the latter ring appear to have a conformation similar to that of the protein molecules in the virus.

Distribution of RNA and protein in crystalline eukaryotic ribosomes

Two low-resolution three-dimensional maps of the structure of crystalline ribosomes from the oocytes of the lizard, Lacerta sicula, have been obtained by electron microscopy and image processing. One map, derived from sheets contrasted with gold-thioglucose, shows the whole ribosome in outline. The other map, based on sheets embedded in glucose, shows predominantly the RNA in the ribosome. The distribution of RNA-rich and protein-rich regions within the ribosome was assessed by comparing both maps. The RNA forms a dense central core, while the ribosomal protein is located mainly at the periphery and constitutes most of the ribosome surface. The RNA appears to be accessible at several sites on the surface. The two subunits of the ribosome are not resolved, indicating that they are in close contact with one another. The subunit interface cuts through a region of the ribosome that is particularly rich in RNA.

Three-dimensional model of membrane-bound ribosomes obtained by electron microscopy

A low-resolution three-dimensional map has been obtained from crystalline arrays of membrane-bound eukaryotic ribosomes. It shows both ribosomal subunits to be adjacent to the membrane surface, attached to it by a part protruding from the large subunit.