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The pyruvate dehydrogenase multienzyme complex of Bacillus stearothermophilus was treated with chymotrypsin at pH 7 and 0 degrees C. Loss of the overall catalytic activity lagged behind the rapid cleavage of the lipoate acetyltransferase polypeptide chains, whose apparent Mr fell from 57 000 to 45 000 as judged by sodium dodecylsulphate/polyacrylamide gel electrophoresis. The inactive chymotrypsin-treated enzyme had lost the lipoic-acid-containing regions of the lipoate acetyltransferase chains, yet remained a highly assembled structure. Treatment of this chymotryptic core complex with trypsin at pH 7.0 and 0 degrees C caused a further shortening of the lipoate acetyltransferase polypeptide chains to an apparent Mr of 28 000 and was accompanied by disassembly of the complex. The lipoic-acid-containing regions are therefore likely to be physically exposed in the intact complex, protruding from the structural core formed by the lipoate acetyltransferase component between the subunits of the other component enzymes. Proton nuclear magnetic resonance spectroscopy demonstrated that the enzyme complex contains large regions of polypeptide chain with remarkable intramolecular mobility, most of which were retained after excision of the lipoic-acid-containing regions with chymotrypsin. It is likely that the highly mobile regions are in the lipoate acetyltransferase component and facilitate movement of the lipoic acid residues. Such polypeptide chain mobility provides the molecular basis of a novel system of active-site coupling in the 2-oxo acid dehydrogenase multienzyme complexes.

To examine the effect of contralateral carotid artery stenosis on postoperative events, a retrospective review was made of 451 patients undergoing 510 carotid endarterectomies during a 6-year period. Three degrees of contralateral carotid stenosis were identified radiologically: 0% to 49%, 50% to 99%, and totally occluded. Each group was further separated into two categories according to preoperative symptoms. "Low risk" included asymptomatic lesions, transient ischemic attacks, and nonhemispheric symptoms; "high risk" described poststroke patients and urgent operations. The results show the incidence of stroke or death was not increased in patients with severely stenosed or occluded contralateral vessels in either low- or high-risk patients (p = 0.741 and p = 0.561, respectively). Patients in the high-risk category, however, had a significantly higher risk of postoperative complications than patients in the low-risk category (p less than 0.001). The study reaffirms that preoperative indications have a major influence on surgical outcome and suggests that the status of the contralateral artery has little bearing on postoperative events.

A comparison is made between the electron density maps of the monoclinic and orthorhombic crystal forms of yeast tRNA Phe which have been obtained respectively by ourselves and by another group. It is concluded that the molecular structures are essentially the same in both crystals, although the models derived from the maps are not the same. The relation between the two molecular packings is discussed, and it is suggested that the intermolecular contact in the orthorhombic form which is not present in the monoclinic form, may arise through base pairing of the anticodons of neighboring molecules.

Subgenomic RNAs (sgRNAs) were isolated from defective interfering virus produced by high multiplicity passage of the human influenza strain A/PR/8/34. Cloning and sequencing of 35 unique sgRNAs revealed that many were about 400 nucleotides long, containing about 200 nucleotides from each of the 5′ and 3′ ends of a full-length segment. Most of the sgRNAs were derived from segment 1, but there were examples from six other segments, including those encoding the haemagglutinin and neuraminidase. Our analysis of the sequence rearrangements found in sgRNAs indicates that they may be generated from the standard viral segments by a jumping viral polymerase that makes transitions between adjacent regions of the RNA template in the ribonucleoprotein tertiary structure.

The nucleosome core particle may be regarded as a basic structural unit of chromatin and thus of the chromosome. It is obtained by micrococcal nuclease digestion of chromatin and consists of about equal weights of DNA (146 base pairs) and protein (an octamer of two pairs each of the histones H2A, H2B, H3, and H4). Nucleosome structure has been reviewed by McGhee and Felsenfeld (6). Studies by small-angle neutron scattering using D2O/H2O contrast variation have given the overall dimensions of the nucleosome and have shown that the protein forms an inner core with the DNA located at the outer regions of the molecule (7,12). The x-ray crystal structure of the nucleosome core particle has been solved in projection to a resolution of about 25 A (3). Although this has given a more detailed description of the overall shape of the molecule than has solution scattering, the distinction between protein and DNA is not made. In this paper we show how the principle of contrast variation can be used in low resolution neutron diffraction to achieve this end.

Summary The three-dimensional reconstruction from electron micrographs of a virus isolated from larvae of the pine emperor moth, Nudaurelia capensis β virus, shows that its surface structure is based on the T = 4 icosahedral surface lattice. The 240 protein subunits are clustered in trimers with the four trimers per icosahedral face approximately planar but with deep grooves between adjacent faces.

Poly(dA).poly(dT) has unusual properties in that it cannot associate into nucleosomes and short, phased runs of it cause DNA bending. The crystal structure of a B-type DNA dodecamer containing a homopolymeric run of six A.T base pairs shows that this region possesses special structural features, including a system of bifurcated hydrogen bonds, which explains some of the properties of this simple homopolymer.

The higher-order structure of chromatin isolated from sea urchin sperm, which has a long nucleosomal DNA repeat length (approximately 240 bp), has been studied by electron microscopy and X-ray diffraction. Electron micrographs show that this chromatin forms 300 A filaments which are indistinguishable from those of chicken erythrocytes (approximately 212 bp repeat); X-ray diffraction patterns from partially oriented samples show that the edge-to-edge packing of nucleosomes in the direction of the 300 A filament axis, and the radial disposition of nucleosomes around it, are both similar to those of the chicken erythrocyte 300 A filament, which is described by the solenoid model. The invariance of the structure with increased linker DNA length is inconsistent with many other models proposed for the 300 A filament and, furthermore, means that the linker DNA must be bent. The low-angle X-ray scattering in the 300-400 A region both in vitro and in vivo differs from that of chicken erythrocyte chromatin. The nature of the difference suggests that 300 A filaments in sea urchin sperm in vivo are packed so tightly together that electron-density contrast between individual filaments is lost; this is consistent with electron micrographs of the chromatin in vitro.

A study of undergraduate medical students' abilities to identify salient information in reviewing patient charts was conducted by members of the Department of Surgery at Southern Illinois University School of Medicine. Specific goals of the two-year study were to develop and test a method for assessing chart skills and to test the following hypotheses: (a) knowledge base is a component of chart review skill; (b) chart skills are related to basic observational skills; (c) performance on one chart is positively correlated with performance on other charts; (d) chart performance is affected by distraction and time pressure; and (e) chart performance improves with clinical experience. Evaluation instruments and methodology are described. Hypotheses a, b, and e and the time effect hypothesized in d were supported; hypothesis c was not supported. Results of the study provide a foundation for further research exploring the effect of instruction on chart performance and investigating the relationships between chart performance and other measures of clinical competence.

Earlier papers in this series on the electron microscopy of negatively-stained human and rabbit papilloma viruses have shown both particles to be composed of 72 morphological units arranged on the T = 7 icosahedral surface lattice. This surface lattice is skew and can exist in either right- or left-handed forms: the hand of the human virus is dextro ( T = 7 d ) and that of the rabbit virus is laevo ( T = 7 l ). The absolute hand of the structure was determined in each case from images of particles which were stained dominantly on one side, but these “one-side” images form only a small proportion of the total. This paper presents the results of experiments in which “two-side” electron microscope images of the same virus particles obtained before and after tilting the supporting grid are compared with simulated images, i.e. superposition patterns from an appropriate model tilted through the same angle. The electron microscope images used were taken from special fields of view—close-packed arrays of particles in a sheet of negative stain—in which the particle structure is well preserved. The superposition patterns were computed and photographed from a display screen, the parameters of the model being varied until the detail in certain distinctive patterns agreed best with that in the corresponding electron microscope images (Plate I). Once the best model was obtained, a gallery of superposition patterns was calculated as a function of the spherical co-ordinates of the axis of view (e.g. Plate IV). When two superposition patterns are selected from the gallery which individually give the best fit for the pair of electron microscope images obtained from the same virus particle before and after tilting the grid, the axis and the angle of rotation to get from one pattern to the other, is, within experimental error, exactly the same as the axis and angle of tilt used in the experiment (Plate V). Moreover, the observed sense of rotation is consistent only with a dextro structure for the human virus and the laevo structure for the rabbit virus. There is very good correlation in detail between the electron microscope images and the simulated images computed for the model. This correspondence extends right to the periphery of the computed superposition patterns (Plate VI) and shows the faint outermost detail in the images to be meaningful, attesting to the faithfulness with which the negative stain preserves and outlines the structure. In the Appendix, the symmetry relationships between the superposition patterns obtained along different axes of view are discussed with the aid of a stereogram.

One group of small DNA-containing animal parvoviruses has already been dealt with in a recent chapter (Rose, this series, Volume 4). The two groups of viruses which come next in size, the polyoma viruses and papilloma viruses, have attracted a great deal of attention especially in recent years. The properties and suggested nomenclature for viruses in these groups are covered in a recent publication (Melnick et al., 1974). In this chapter we will review the present state of knowledge regarding the chemical and physical make-up of these viruses.

Publisher Summary This chapter describes the X-ray crystallographic and enzymatic analysis of nucleosome cores. Almost all preparations of rat liver nucleosome cores under the appropriate conditions yielded crystalline material provided the material was homogeneous enough. Crystallization of nucleosome cores has been reported. It is found that while intact nucleosome cores have been crystallized and studied, one have found that the largest crystals so far obtained, which diffract to spacings of about 20 A, come from particles which had some of their histones cleaved by endogenous proteolysis. The sedimentation constant of the proteolysed particle in 100 mM NaCl is hardly changed from the intact material. Brief DNase I digestion shows a pattern of bands every 10 bases which is very similar in its intensity distribution to that from intact particles, except that the rate of digestion of the proteolysed particles is higher. The relative frequencies of cutting at the individual sites, as determined by end-labeling experiments, remain very similar for the proteolysed particles.

Methods are developed for computing three dimensional density maps from transmission electron micrographs using Fourier transforms. The reconstructions objectively combine data from several different views of one or more particles. When applied to negatively stained spherical viruses, the technique gives clear reconstructions of their surface structures.

A comparative study of X-ray powder diagramsindicates that the structure of the RNA in both microsomal particles and small viruses is very different from that of isolated RNA. The conclusion is that in these particles the RNA is not segregated, but takes on a mole-cular configuration determined by the form of the protein matrix

The hydrous ferric oxide (Fe2O3 · xH2O) catalyst developed by Loebenstein and Weitzel[1] has been shown to be quite effective for ortho-para hydrogen conversion. The desire to obtain maximum activity for the hydrous ferric oxide catalyst has led to various activation procedures by previous investigators[2,3,4]. These methods consisted of holding the catalyst at an elevated temperature while either pumping to a moderate vacuum or purging with a pure gas for some specified length of time.

VARIOUS species of transfer RNA have been crystallized by controlled precipitation from aqueous solutions containing organic solvents or ammonium sulphate (reviewed in refs. 1 and 2). These methods have produced a great variety of crystal forms which, with a few exceptions3,4, are usually poorly ordered as judged by X-ray diffraction. This is probably because the interactions between molecules are few and rather nonspecific, making the crystal structure extremely sensitive to the crystallization conditions. For this reason, attempts have been made to crystallize tRNA as the cetyltrimethylammonium (CTA-) salt. The additional interaction between hydrophobic cetyl cations bound to the different molecules may stabilize the crystal lattice and have a positive effect on the crystallization process and therefore on the order of the crystals. We report here the production of crystals of CTA-salts of five different tRNAs; tRNAMetf, tRNAGlu, tRNAPhe, tRNATyr from E. coli and tRNAPhe from yeast. In the case of tRNAMetf, different crystal forms were obtained in the presence of different cations.

X-ray diffraction studies of crystals of turnip yellow mosaic virus and of the associated protein particles have led to new results concerning (a) the packing of the virus particles in the crystal cell, and (b) the arrangement of sub-units in a virus particle. In the virus crystals the particles lie with their centres at the lattice points of a body-centred cubic unit cell of side 350 A, but are not all in the same orientation so that the true crystallographic cell has a side of 700 A and contains 16 particles. The virus particle as a whole has point-group symmetry 23, and is therefore built up of 12, or a multiple of 12, sub-units. The additional symmetry of the point-group 532 is also partially present. It is concluded that there are probably 60 sub-units forming the protein “shell” of the virus.

Self-assembly of the protein subunits of tobacco mosaic virus takes place by a series of steps. The natural state of aggregation of TMV protein is not the helix but the 20S disk, which consists of two paired rings of seventeen sub-units each. Movement of 5 A is involved in the pairing. Control of the aggregation and its consequences for virus assembly are described in the next two articles.

Summary Several plant viruses with filamentous particles ranging in modal lengths from 0.48 µ to 1.25 µ were negatively stained with uranyl formate, examined in the electron microscope, and the electron micrographs analysed in various ways. The particles of all the viruses were helically constructed with a basic pitch of 33 to 37 A (mean 34 A), but could be separated into groups by other features of their particles. Various measurements of the particles of five of the viruses suggest that there were 10 to 14 subunits in each turn of the basic helix of their particles. All plant viruses with elongated particles seem to fall into one of two groups; those with modal lengths of 0.3 µ or less seem rigid and have a basic helix of pitch 23 to 25 A, and those with longer particles are filamentous and have a basic helix of pitch 33 to 37 A.