A new approach to improving the accuracy of elemental analysis in laser mass spectrometry.

It is shown that the main reason affecting the accuracy and reproducibility of the elemental analysis of solids with a laser source of ions is the difference in the ionization cross sections of different elements in the generation of laser plasma. Computer modeling was carried out for the evaporation of the sample and generation of laser plasma at different values of laser power density. The aim of modelling was to determine the cause of the low accuracy of analysis by direct selection of ions from the plasma. The researches have shown that we cannot get satisfactory accuracy of analysis using analytical signal formed only on the base of single-charged ions. At the same time, in the formation of an analytical signal, as a sum of the intensities of the mass peaks of all charges of each element, the accuracy of the analysis does not depend either on the ionization cross section or on the nonreproducibility of the laser radiation power. It is shown that this approach completely eliminates the "matrix effect."

Expression, purification and crystallization of a thermostable short-chain alcohol dehydrogenase from the archaeon Thermococcus sibiricus.

Alcohol dehydrogenases belong to the oxidoreductase family and play an important role in a broad range of physiological processes. They catalyze the cofactor-dependent reversible oxidation of alcohols to the corresponding aldehydes or ketones. The NADP-dependent short-chain alcohol dehydrogenase TsAdh319 from the thermophilic archaeon Thermococcus sibiricus was overexpressed, purified and crystallized. Crystals were obtained using the hanging-drop vapour-diffusion method using 25%(w/v) polyethylene glycol 3350 pH 7.5 as precipitant. The crystals diffracted to 1.68 A resolution and belonged to space group I222, with unit-cell parameters a = 55.63, b = 83.25, c = 120.75 A.

Crystallization and preliminary X-ray diffraction analysis of Salmonella typhimurium uridine phosphorylase complexed with 5-fluorouracil.

Uridine phosphorylase (UPh; EC 2.4.2.3) catalyzes the phosphorolytic cleavage of the N-glycosidic bond of uridine to form ribose 1-phosphate and uracil. This enzyme also activates pyrimidine-containing drugs, including 5-fluorouracil (5-FU). In order to better understand the mechanism of the enzyme-drug interaction, the complex of Salmonella typhimurium UPh with 5-FU was cocrystallized using the hanging-drop vapour-diffusion method at 294 K. X-ray diffraction data were collected to 2.2 A resolution. Analysis of these data revealed that the crystal belonged to space group C2, with unit-cell parameters a = 158.26, b = 93.04, c = 149.87 A, alpha = gamma = 90, beta = 90.65 degrees . The solvent content was 45.85% assuming the presence of six hexameric molecules of the complex in the unit cell.

Crystallization and preliminary x-ray diffraction study of neurotoxin-I from Naja naja oxiana venom.

Crystals of the neurotoxin-I (NTX-I) from the venom of the middle Asian cobra Naja naja oxiana have been grown by vapour diffusion and dialysis methods. The crystals belong to space group P2(1)2(1)2 with dimension of a = 25.19 A, b = 75.59 A, c = 36.09 A and diffract to 1.9 A resolution. The asymmetric unit contains one molecule (Vm = 2.2 A/Da). Using the molecule of alpha-cobratoxin (CTX) as a starting model for NTX-I structure determination coordinates of C alpha atoms of the NTX-I molecule were obtained and the position of NTX-I in the unit cell was derived.

The atomic structure of Carnation Mottle Virus capsid protein.

The structure of the Carnation Mottle Virus (CMtV) capsid protein has been determined at 3.2 A resolution by the method of molecular replacement. Three-dimensional data were collected from a small number of crystals (sp.g. I23, a = 382.6 A) using the synchrotron radiation with an image plate as detector. The coordinates of Tomato Bushy Stunt Virus (TBSV) were used as a searching model. Refinement of the coordinates of 7,479 non-hydrogen atoms performed by the program XPLOR, has led to an R-factor of 18.3%. It was found that the amino acid chain fold of capsid protein is very similar to that in other icosahedral viruses. However, there are some differences in the contact regions between protein subunits and also the lack of the beta-annulus around the 3-fold icosahedral axes. The structural and biochemical results lead us to consider an alternative assembly pathway.

Three-dimensional structure of Serratia marcescens nuclease at 1.7 A resolution and mechanism of its action.

The three-dimensional crystal structure of Serratia marcescens (Sm) nuclease has been refined at 1.7 A resolution to the R-factor of 17.3% and R-free of 22.2%. The final model consists of 3678 non-hydrogen atoms and 443 water molecules. The analysis of the secondary and the tertiary structures of the Sm nuclease suggests a topology which reveals essential inner symmetry in all the three layers forming the monomer. We propose the plausible mechanism of its action based on a concerted participation of the catalytically important amino acid residues of the enzyme active site.

Atomic structure at 2.5 A resolution of uridine phosphorylase from E. coli as refined in the monoclinic crystal lattice.

Uridine phosphorylase from E. coli (Upase) has been crystallized using vapor diffusion technique in a new monoclinic crystal form. The structure was determined by the molecular replacement method at 2.5 A resolution. The coordinates of the trigonal crystal form were used as a starting model and the refinement by the program XPLOR led to the R-factor of 18.6%. The amino acid fold of the protein was found to be the same as that in the trigonal crystals. The positions of flexible regions were refined. The conclusion about the involvement in the active site is in good agreement with the results of the biochemical experiments.

[Spatial structure of the basic chain of the neurotoxin I molecule from the venom of cobra Naja naja oxiana and its crystal packing]. / Prostranstvennoe stroenie osnovnoi tsepi molekuly neirotoksina I iz iada kobry Naja naja oxiana i ee ukladka v kristallicheskoi iacheike.

The structure of the alpha-carbon chain was solved by molecular replacement method at 2.7 A resolution. Neurotoxin I (NTX-I) is one of the main protein components purified from the venom of the central asian cobra Naja naja oxiana. NTX-I is known to bind specifically to acetylcholine receptors thus preventing the transmission of the neuroconductivity signal from synapses to muscles. NTX-I crystals were grown either by vapour diffusion or dialysis methods using specially prepared microdialysis cell. The intensities of reflections from native NTX-I crystals were measured in the range of 38.0-2.1 A-1 by omega-scan method with a Syntex P21 diffractometer operated in automatic regime. To determine the position and mode of packing of NTX-I molecule in unit cell program packages MERLOT and BLANC were applied running on a NORD-500 computer.

[X-ray structural analysis of magnesium-containing Serratia marcescens endonuclease]. / Rentgenostrukturnyi analiz magniisoderzhashchei éndonukleazy Serratia marcescens.

The three-dimensional crystal structure of the DNA/RNA nonspecific endonuclease from Serratia marcescens was refined at the resolution of 1.07 A to R factor of 12.4% and Rfree factor of 15.3% using the anisotropic approximation. The structure includes 3924 non-hydrogen atoms, 715 protein-bound water molecules, and a Mg2+ ion in each binding site of each subunit of the nuclease homodimeric globular molecule. The 3D topological model of the enzyme was revealed, the inner symmetry of the monomers in its N- and C-termini was found, and the local environment of the magnesium cofactor in the nuclease active site was defined. Mg2+ ion was found to be bound to the Asn119 residue and surrounded by five associated water molecules that form an octahedral configuration. The coordination distances for the water molecules and the O delta 1 atom of Asn119 were shown to be within a range of 2.01-2.11 A. The thermal factors for the magnesium ion in subunits are 7.08 and 4.60 A2, and the average thermal factors for the surrounding water molecules are 11.14 and 10.30 A2, respectively. The region of the nuclease subunit interactions was localized, and the alternative side chain conformations were defined for 51 amino acid residues of the nuclease dimer.

[A comparative structure-function analysis and molecular mechanism of action of endonucleases from Serratia marcescens and Physarum polycephalum]. / Sravnitel'nyi strukturno-funktsional'nyi analiz éndonukleaz Serratia marcescens i Physarum polycephalum i molekuliarnyi mekhanizm ikh deistviia.

Structural and functional characteristics were compared for wild-type nuclease from Serratia marcescens, which belongs to the family of DNA/RNA nonspecific endonucleases, its mutational forms, and the nuclease I-PpoI from Physarum polycephalum, which is a representative of the Cys-His box-containing subgroup of the superfamily of extremely specific intron-encoded homing DNases. Despite the lack of sequence homology and the overall different topology of the Serratia marcescens and I-PpoI nucleases, their active sites have a remarkable structural similarity. Both of them have a unique magnesium atom in the active site, which is a part of the coordinatively bonded water-magnesium complex involved in their catalytic acts. In the enzyme-substrate complexes, the Mg2+ ion is chelated by an Asp residue, coordinates two oxygen atoms of DNA, and stabilizes the transition state of the phosphate anion and 3'-OH group of the leaving nucleotide. A new mechanism of the phosphodiester bond cleavage, which is common for the Serratia marcescens and I-PpoI nucleases and differs from the known functioning mechanism of the restriction and homing endonucleases, was proposed. It presumes a His residue as a general base for the activation of a non-cluster water molecule at the nucleophilic in line displacement of the 3'-leaving group. A strained metalloenzyme-substrate complex is formed during hydrolysis and relaxes to the initial state after the reaction. The English version of the paper.

[Three-dimensional reconstruction of bacterial viruses from electron microscopy data. Bacteriophage structure]. / Trekhmernaia rekonstruktsiia bakterial'nykh virusov po dannym élektronnoi mikroskopii. Stroenie bakteriofagov.

The structure of bacteriophages from different groups is presented based on the data obtained by the three-dimensional reconstruction, optical diffraction and filtration and electron microscopy techniques. The study made possible to suggest the scheme for the mechanism of sheath molecules rearrangement at contraction of bacteriophage tail sheath, the model of bacteriophage FI-1 connector. The structural elements for difference and relation of various bacteriophages are demonstrated.

[Three-dimensional reconstruction of bacterial viruses from electron microscopy data. Subjects and methods of study]. / Trekhmernaia rekonstruktsiia bakterial'nykh virusov po dannym élektronnoi mikroskopii. Ob''ekty i metody issledovaniia.

The review summarizes the results of the study of the spatial structure of bacterial viruses (phages) whose tails seem to be the most primitive contracting biological mechanism. Data on the spatial molecular rearrangement are important for understanding the processes of biological mobility. The computer and laser techniques used in order to obtain information on the three-dimensional structure of the object under study by its two-dimensional electron-microphotography are presented in the first part of the review. The second deals with application of the above mentioned techniques for the study of various bacterial viruses.

[The submolecular structure of the Carnation Mottle virus]. / O submolekuliarnom stroenii viriona Carnation Mottle Virus.

The model of the tertiary structure of the CMtV subunit was suggested and the mode of its packaging into the capsid was determined on the basis of electron microscopy small-angle X-ray scattering and X-ray diffraction data. The i-ray diffraction data on the crystal of the native virion of Carnation Mottle Virus were obtained on the automatic diffractometer supplied with two-dimensional detector.

[Viscumin modulates neutrophil respiratory burst, caused by a chemotactic peptide]. / Viskumin moduliruet respiratornyi vzryv v neitrofilakh, vyzvannyi khemotaksicheskim peptidom.

The ability of viscum at different concentrations to modulate the respiratory burst in neutrophils, induced by the chemotactic peptide N-formyl-methionyl-leucyl-phenylalanine was studied. This does not exclude the possibility that viscum can interact with the receptor of this peptide. The analysis of the primary structure of viscum revealed elements structurally analogous to the chemotactic peptide. It is assumed that viscum can exhibit the properties an antagonist of the receptor of N-formyl-methionyl-leucyl-phenylalanine, and the mechanism of action of viscum depends on its concentration.

[Features of the modulating effect of complexes of ribosome-inactivating proteins type II with sugars on respiratory burst of neturophils, caused by a chemotactic peptide]. / Osobennosti moduliruiushchego deistviia kompleksov ribosom-inaktiviruiushchikh belkov II tipa s sakharami na respiratornyi vzryv neitrofilov, vyzvannyi khemotaksicheskim peptidom.

It was shown that agents inducing phagocytosis (zymosan, lectins) cause changes in the number of receptors responsible for fast neutrophil reaction (chemotaxis or respiratory burst) or inhibit the binding of the agonist to its receptor. Among lectins are ribosome-inactivating proteins of type II ricin and agglutinin ricin, which penetrate the cell by binding to mannose and galactose receptors. It was shown that ribosome-inactivating proteins of type II can exhibit the properties of the antagonist of the receptor N-formylmethionylleucylphenylalanine. Ricin is more effective in modulating the respiratory burst induced by the chemotactic peptide than agglutinin ricin. The modulating effect of ribosome-inactivating proteins of type II on neutrophils is likely to be mediated by their interaction with galactose rather than mannose receptors. Presumably, the affinity of ribosome-inactivating proteins to galactose receptors increases with increasing amount of saccharides bound to the protein molecule. The modulating effect of ribosome-inactivating proteins of type II on the respiratory burst of neutrophils induced the chemotactic peptide is due to the structural peculiarities of these proteins.

[The structure of cholera phages 493 and 7227]. / Stroenie kholernykh fagov 493 i 7227.

Electron microscopy and optic diffraction methods were used to determine the main parameters of the morpho-structural organization of cholera phages 493 and 7227. The shape of phage capsids was established, the spiral type of symmetry of phage processes revealed, their adsorption apparatuses described, the number of morphological subunits comprising capsids and process envelopes, and some other parameters determined. It is assumed that the phages under study are new varieties of bacteriophages belonging to the fifth morphological group (according to the classification of A.S. Tikhonenko).

[Electron microscopic study of the structure of cholera bacteriophage serotype II]. / Elektronno-mikroskopicheskoe issledovanie struktury kholernogo bakteriofaga II serotipa.

The methods of electron microscopy and optic difraction were used to determine the main parameters of the structural organization of cholera bacteriophage of serotype II. The virions of the phage under study were found to consist of a capsid of icosahedral shape and 77.4 X 66.1 nm in size, formed of 252 morphological subunits and having the cavity volume of 76.3 X 10(3) nm3. A process of 94.8 nm in length adheres to the capsid, with a shell capable of contractions, of 85.6 nm in length, 20.6 nm in width, containing 60 morphological subunits. The shell covers the central core of 8.9 nm in diameter. The optic difraction revealed the helical type of symmetry of the process structure and its main parameters.

[Purification of angiogenin from cow's milk]. / Ochistka angiogenina iz korov'evo moloka.

A new method for the purification of bovine angiogenin is proposed which is based on the use of CM-cellulose, CM-Toyopearl, and Butyl-Toyopearl. Electrophoretically homogeneous protein was obtained with a 49% yield and 12,000-fold purification.

[Atherogenesis in bronchial asthma patients: its relationship to the pathogenetic characteristics of the disease and to liver function (the clinico-biochemical aspects)]. / Aterogenez u bol'nykh bronkhial'noi astmoi, ego zavisimost' ot patogeneticheskikh osobennostei zabolevaniia i funktsional'nogo sostoianiia pecheni (kliniko-biokhimicheskie aspekty).

Ten asthmatics in remission and aggravation of the disease were examined for: lipoprotein pre-beta, beta and alpha fractions, total cholesterol and its fractions in atherogenic and antiatherogenic lipoproteins with estimation of the atherogenicity coefficient, activity of 5-nucleotide hydrolase, acid phosphatase, gamma-glutamyl transpeptidase and alcohol dehydrogenase. The results were indicative of more active atherosclerotic process in the presence of blockade which changed the activity of hepatocyte membrane lipoprotein receptors, of simultaneous operation in bronchial asthma patients of infiltrative and cellular mechanisms of atherosclerosis.

[Determination of angiogenin in cow's milk based on a competitive test "pancreatic RNAase/placental inhibitor of RNAase--angiogenin"]. / Opredelenie angiogenina v korov'em moloke na osnove konkurentnoi proby "pankreaticheskaia RNKaza/platsentarnyi ingibitor RNKazy-- angiogenin".

Contents of active factor of blood vessel growth angiogenin was analyzed in cow's milk by the method of competitive test in system "pancreatic RNAase/placental inhibitor of RNAase-angiogenin". High level of RNAase in milk limits using this test. To avoid this limitation the method of RNAase elimination from milk was elaborated.