Preparative method of isolating phosphoinositide fractions from brain tissue]

A simple technique for preparative isolation of chromatographically homogenous fractions of mono-, di- and triphosphoinositides from ox brain tissue is described. Podyphosphoinositides fractions were obtained after chromatography of lipid extract on DEAE cellulose, phosphomonoinositides fraction--after chromatography of polyphosphoinositide-free material on aluminium hydroxide column. Bivalent metal ions were eliminated from lipid extract using chromatography on Dowex-50 H+. Ammonium acetate was removed after precipitation of lipids in water: methanol (1:1) in the presence of 4 M this salt. The average yield of mono-, di- and triphosphates was 40, 22 and 58 mg respectively per 1 kg of brain tissue as callulated for lipid phosphorus.     

Immunomorphological changes in the mouse brain after intracerebral administration of a neurotropic strain of influenza virus]

The work dealt with intracerebral inoculation of 150 mice with neurotropic strain of influenza virus A/WSN/33. Virological and immunohistochemical study of virus localization in the mice brain was carried out. It was shown that virus reproduction in ependymal lining cells and plexus choroideus epithelium is followed by destruction of cells and uptake of disintegration products by macrophages. Since the influenza infection is followed by a large number of macrophages involved in gene expression of the main histocompatibility complex, the development of autoimmune reaction with the formation of autoantibodies to brain specific antigens can't be ruled out.     

An invisible ubiquitin conformation is required for efficient phosphorylation by PINK1

The Ser/Thr protein kinase PINK1 phosphorylates the well‐folded, globular protein ubiquitin (Ub) at a relatively protected site, Ser65. We previously showed that Ser65 phosphorylation results in a conformational change in which Ub adopts a dynamic equilibrium between the known, common Ub conformation and a distinct, second conformation wherein the last β‐strand is retracted to extend the Ser65 loop and shorten the C‐terminal tail. We show using chemical exchange saturation transfer (CEST) nuclear magnetic resonance experiments that a similar, C‐terminally retracted (Ub‐CR) conformation also exists at low population in wild‐type Ub. Point mutations in the moving β5 and neighbouring β‐strands shift the Ub/Ub‐CR equilibrium. This enabled functional studies of the two states, and we show that while the Ub‐CR conformation is defective for conjugation, it demonstrates improved binding to PINK1 through its extended Ser65 loop, and is a superior PINK1 substrate. Together our data suggest that PINK1 utilises a lowly populated yet more suitable Ub‐CR conformation of Ub for efficient phosphorylation. Our findings could be relevant for many kinases that phosphorylate residues in folded protein domains.     

Detection of somatic mosaicism in humans using polymerase chain reaction with random primers

Using the method PCR amplification with random primers, DNA samples from human embryonic organs and tissues were examined. Among 27 oligonucleotide primers tested, 10 primers, producing stable, well-reproducible profiles of amplification products, were chosen for further analysis. With the help of two primers (447 and R45), the differences in RAPD PCR profiles obtained from the tissues of one embryo, were revealed. These differences were associated with the change of mobility, or with the fragment gain/loss in the RAPD profile, and could be caused either by genomic rearrangements, or mutations involving the regions of the DNA-primer pairing. Different epigenetic factors, like methylation, can also play the role in this process.     

Preservation of Neisseria genus microbes by a deep freezing method]

A method of meningococcus and of vaccine gonococcus strains preservation under conditions of deep freezing with polyethyleneoxide of mol mass 400 was developed. Peculiarities of phasic transitions were studied by the method of differential thermal analysis; hydration properties of polyethyleneoxide were investigated by nuclear magneticresonance (spin-echo). The main cultural and biochemical properties after freezing and after one year of storage in 8 strains of the microbes under study remained unchanged; their survival constituted not less than 90%.     

The cloning and expression of the Staphylococcus aureus enterotoxin A gene in Escherichia coli cells

Gene ent-A has been cloned on phage vector pSL5 with the use of the gene library of S. aureus FR1722(H). It is located within DNA fragment Hind III having 2,500 nucleotide pairs.     

On the structure of liver ribosomes

The morphology of liver ribosomes and their subparticles, large and small, has been investigated. Analysis of the images has been carried out by successive selection of models and by X-raying them under conditions simulating negative staining. The relation between the main views has been checked by tilting the specimens in an electron microscope through ± 30 °.The small subparticle consists of an elongated body, to one of the ends of which a short “head” is attached. A model has been proposed, whose projections on rotation with respect to the longitudinal axis would satisfy all observable types of images. According to the proposed model, the “head” is tilted with respect to the elongated portion. The length of the subparticle is 230 Å. The dimensions of the elongated portion in the transverse direction are 110 Å × 75 to 80 Å.The large subparticles in lateral view resemble short “rods” 220 to 240 Å long and about 70 to 95 Å wide. At a distance of about 60 Å from the left end of the particles a projection (60 Å in length) is seen, on the inner side of which a depression, or “channel”, filled with the contrasting substance is always observed. Next to this depression a second projection is located, whose height is about 30 Å. The channel is either a cavity in the body of the large subparticle or a part of the RNA without protein. The length of the channel is about 80 Å, the width is about 50 to 60 Å. The left end of the particles is characterized by two sharpened portions; as a result, a cavity that shows an obtuse angle profile makes its appearance. The opposite end of the particles is cut off at an angle of 45 °. In another view, the subparticles appear to be almost rectangular in shape; they are characterized by dimensions of 150 Å × 220 to 240 Å. It is likely that the large projection is displaced sideways with respect to the longitudinal axis of the particles. The asymmetry associated with this displacement gives rise to preferred arrangements of the subparticles on the supporting film. An analysis has been made of the most typical images of monomeric ribosomes, on the basis of which a suggestion is made about mutual orientation of subparticles in a monomer.