Theoretical Study of Antagonists and Inhibitors of Mammalian Adenosine Deaminase: II. Isomeric Aza-deazaanalogues of Adenosine

Isomeric aza-deazaanalogues of adenosine and their N1-protonated forms (except for that of 8-aza-1-deazaadenosine) were studied by computer modeling to find a relationship between their molecular structures and the properties as substrates for the mammalian adenosine deaminase. The atomic charge distribution and maps of electrostatic potential around their van der Waals molecular surface were calculated using the ab initioSTO-3G method. The conformational studies were carried out by the MM⁺ method of molecular mechanics. The previously proposed mechanism of the substrate acceptance in the active site of mammalian adenosine deaminase was refined, and the potential substrate properties were predicted for two previously unstudied adenosine analogues, 5-aza-9-deazaadenosine and 8-aza-3-deazaadenosine.     

Sequence-specific ultrasonic cleavage of DNA

We investigated the phenomenon of ultrasonic cleavage of DNA by analyzing a large set of cleavage patterns of DNA restriction fragments using polyacrylamide gel electrophoresis. The cleavage intensity of individual phosphodiester bonds was found to depend on the nucleotide sequence and the position of the bond with respect to the ends of the fragment. The relative intensities of cleavage of the central phosphodiester bond in 16 dinucleotides and 256 tetranucleotides were determined by multivariate statistical analysis. We observed a remarkable enhancement of the mean values of the relative intensities of cleavage (cleavage rates) in phosphodiester bonds following deoxycytidine, which diminished in the row of dinucleotides: d(CpG) > d(CpA) > d(CpT) >> d(CpC). The cleavage rates for all pairs of complementary dinucleotides were significantly different from each other. The effect of flanking nucleotides in tetranucleotides on cleavage rates of all 16 types of central dinucleotides was also statistically significant. The sequence-dependent ultrasonic cleavage rates of dinucleotides are consistent with reported data on the intensity of the conformational motion of their 5′-deoxyribose. As a measure of local conformational dynamics, cleavage rates may be useful for characterizing functional regions of the genome.     

Theoretical study of antagonists and inhibitors of mammalian adenosine deaminase. I. Adenosine and its aza- and deaza-analogs

Aza- and deazaanalogues of adenosine, including their 1-protonated forms (except for that of 1-deazaadenosine), were studied by computer computation to find a relationship between their molecular structures and substrate properties for the mammalian adenosine deaminase. The atomic charge distribution and maps of the electrostatic potential around their van der Waals molecular surface were calculated for these compounds using the ab initio STO-3G method. The conformational studies were carried out by the MM+ method of molecular mechanics. The mechanism that determines the substrate selectivity of mammalian adenosine deaminase is discussed. The English version of the paper: Russian Journal of Bioorganic Chemistry, 2002, vol. 28, no. 4; see also http://www.maik.ru.     

Absorption and release of biogenic amines by the lung during the restorative period after hypovolemic hypotension

The processes of serotonin and histamine absorption and release by the lungs were studied in dogs during 1 to 3.5 hour hypovolemic hypotension and during 24 hours after blood retains fusion. Absorption of biogenic amines by the lungs tended to increase in all the animals under hypovolemic hypotension. In the group of non-survivors the serotonin absorption by the lungs in the post-terminal period remained increased, while in the group of survivors it came down to normal soon, though the histamine release was increased. The above processes were aggravated in the group of animals whose lungs were affected by oleic acid. It resulted in the absorption of histamine instead of its release. The intensified absorption of biogenic amines by the lungs was accompanied by a quick fall in cardiac output, by the increase in resistance of systemic and pulmonary circulation.     

Characteristics of rat ceruloplasmin from the serum of animals, which received salts of silver with food

Abiogenic Ag(I) ions have electronic structure, similar to Cu(I) ions and can compete with Cu(I) for binding sites of proteins which transport copper from extracellular media to sites of cuproenzyme formation in the cell. Rodents receiving Ag-salts with food develop extracellular deficiency of copper associated with ceruloplasmin (Cp, the major copper-transporting protein in blood serum of vertebrates). The present work focuses on the studies of biochemical and physicochemical properties of Cp, obtained from blood serum of rats, which received AgCl with food for 4 weeks (Ag-rats). Cp-fractions from blood serum of Ag-rats (Ag-Cp) were obtained by ion-exchange chromatography with stepped gradient of NaCl. Each fraction was tested for oxidase and ferroxidase activities by direct measurement of catalytic activity in the gel, and for specific activity in holo-Cp in oxidation of chromogenic substrate. Molecular mass, electrophoretic mobility and ratio of apo- and holo-forms in Ag-Cp fractions were evaluated by immunoblotting. Ag-Cp samples did not contain products of spontaneous partial proteolytic degradation, characteristic of holo-Cp samples. Fractions of Ag-Cp and holo-Cp (from blood serum of control rats) were compared by optical spectra, tertiary structure, susceptibility to thermal denaturation, and by atomic Cu and Ag content. Ag-Cp contained 1-2% Cp, which is similar by spectral and catalytic properties with holo-Cp. [Ag]:[Cu] ration in Ag-Cp samples was about 4:1. As evidenced by circular dichroism and differential scanning calorimetric studies, the major apo-fraction of Ag-Cp lacked tertiary structure of native Cp and was significantly misfolded, which might explain its resistance to spontaneous partial proteolytic degradation.     

Heterogeneity of double-stranded DNA local structure and dynamics: ultrasound studies

A method for studying the local inhomogeneities of DNA and its dynamics is proposed. The method is based on the combination of two procedures, splitting the DNA molecules by ultrasound and analysis of DNA fragments obtained by gel electrophoresis. The frequency of cleavage of internucleotide bonds was found to depend on the type of nucleotides forming the bond and on their nearest neighbors. Estimates of cleavage frequencies in each of 16 dinucleotides showed that, in the 5'-d(CpG)-3', 5'-d(CpA)-3', and 5'-d(CpT)-3' dimers, the cleavage occurs considerably more frequently than in the rest, and the frequency of cleavage depends on the nearest neighbors. It was shown that the double-helix cleavage can occur with shifts by several nucleotides. Physical prerequisites were considered that can lead to this pattern of sequence - specific cleavage.     

Immunostimulating effect of plant triterpenes and their derivatives

The immunostimulating activity of triterpenes of vegetable origin and their derivatives was studied. The study revealed that glycyrrhizic and betulonic acids, as well as derivatives of ursolic and glycyrrhetinic acids, enhanced the level of specific antibodies to bovine serum albumin in mice.     

Conformational Polymorphism and Extensibility of DNA Quadruplexes Formed by d(GT)nRepeats

We showed earlier that oligonucleotides 3"-d(GT)₅-pO(CH₂CH₂O)₃p-d(GT)₅-3" form bimolecular quadruplexes with parallel orientation of their strands, which are held by guanine quartets alternating with unpaired thymines (GT quadruplex). This work deals with the conformational polymorphism and extensibility of G quadruplexes in complex with molecules of an intercalating agent ethidium bromide (EtBr). A cooperative mechanism of EtBr binding to the GT quadruplex was revealed. The binding constant K= (3.3 ± 0.1)·10⁴M^–1, cooperativity coefficient = 2.5 ± 0.2, and maximal amount of EtBr molecules intercalated in GT quadruplex (N= 8) were determined. It was proved experimentally by analysis of adsorption isotherms and theoretically by mathematical modeling that the GT quadruplex is capable of double extension, which is indicative of the high elasticity of this four-stranded helix. Two most stable conformations of GT quadruplexes with thymine residues intercalated and/or turned outside were found by mechanico-mathematical modeling. The equilibrium is shifted toward the conformation with the looped out thymine residues upon intercalation of EtBr molecules into the GT quadruplex.     

Parallel double DNA spiral. A conformational analysis of regular spirals of poly(dA).poly(dT) with different variations of joining bases

We have performed a conformational analysis of DNA double helices poly(dA).poly(dT) with parallel directed backbone strands in heteronomic model frames. All possible models of base pairs and various mutual orientation of base pair and sugarphosphate backbones were checked. By the potential energy optimization the dihedral angles and helices parameters of stable conformations of parallel double polynucleotides were calculated. The dependences of conformational energy on the base pair structure were studied.