[Investigation of the influence of the melanoidin formation reaction on the content of amino acids in model food systems].

Almost all currently used technologies for the food production are related to the melanoidinformation reaction, which has a significant effect on appearance, taste, nutritional value and consumer properties of the foodstuffs. To assess the effect of heat treatment of food products on their nutritional value, food model systems protein hydrolyzate - glucose, hydrolyzate protein - xylose, hydrolyzate protein - fructose (1:5) have been investigated. The influence of the presence of reducing sugars, the temperature and the duration of heating on the content of amino acids and the extinction of solutions of model food systems have been studied. Linear dependences of the decrease in the total amino acid content in model food systems on the duration of the melanoidin formation reaction have been found. The loss of the total amino acid content when heating to 120 °C for 120 min was 23.9%; at the same time the content of essential amino acids reduced by 15.5-24.6%. The addition of xylose intensified the process of destruction of amino acids in the model system by 12.7%, at the same time glucose provoked the destruction of amino acids by only 2.3%. It has been established that amino acids threonine, isoleucine and histidine were unstable to destruction, regardless of the type of added sugar. When white wheat bread was heated, the loss of its nutritional value was established by reducing the content of such essential acids as threonine (by 26.5%), methionine (by 21.2%), lysine (by 13.3%) and valine (by 12,1%). It was noted that at the same temperature with increasing time of the melanoidin formation reaction, the content of amino acids in the system decreased with simultaneous intensification of the staining of the solutions. Extinction of model food systems varied according to the following equations: hydrolyzate-glucose - y = 0.0022x, hydrolyzate-xylose - y = 0.0028x, hydrolyzate-fructose - y = 0.0032x.

Spatial invasion dynamics on random and unstructured meshes: implications for heterogeneous tumor populations.

In this work we discuss a spatial evolutionary model for a heterogeneous cancer cell population. We consider the gain-of-function mutations that not only change the fitness potential of the mutant phenotypes against normal background cells but may also increase the relative motility of the mutant cells. The spatial modeling is implemented as a stochastic evolutionary system on a structured grid (a lattice, with random neighborhoods, which is not necessarily bi-directional) or on a two-dimensional unstructured mesh, i.e. a bi-directional graph with random numbers of neighbors. We present a computational approach to investigate the fixation probability of mutants in these spatial models. Additionally, we examine the effect of the migration potential on the spatial dynamics of mutants on unstructured meshes. Our results suggest that the probability of fixation is negatively correlated with the width of the distribution of the neighborhood size. Also, the fixation probability increases given a migration potential for mutants. We find that the fixation probability (of advantaged, disadvantaged and neutral mutants) on unstructured meshes is relatively smaller than the corresponding results on regular grids. More importantly, in the case of neutral mutants the introduction of a migration potential has a critical effect on the fixation probability and increases this by orders of magnitude. Further, we examine the effect of boundaries and as intuitively expected, the fixation probability is smaller on the boundary of regular grids when compared to its value in the bulk. Based on these computational results, we speculate on possible better therapeutic strategies that may delay tumor progression to some extent.

[Evaluation of compensation abilities of kidneys using radionuclide examination].

The article presents the results of evaluation of compensation abilities of kidney based on the identification of volume of functioning renal tissue according to the single-photon emission scintigraphy (SPECT) and planar scintigraphy. The study included adult patients (65 patients with renal cell carcinoma, 32 patients with renal cysts and 20 patients without renal involvement) and pediatric patients aged 1 to 17 years (57 patients with various non-neoplastic lesions of the kidney). The following parameters were used: volume ratio of functioning tissue and the specific activities of the affected and contralateral kidneys, and the volumetric ratio of volumes estimated to the reference kidney volume depending on anthropomorphic data of patient (age and sex). It was found that the maximum compensatory renal reserve occurs in the case of reducing the volume of functioning tissue of the affected kidney and the simultaneous increase in the volume ofcontralateral kidney.

Engineering of substrate specificity of D-amino acid oxidase from the yeast Trigonopsis variabilis: directed mutagenesis of Phe258 residue.

Natural D-amino acid oxidases (DAAO) are not suitable for selective determination of D-amino acids due to their broad substrate specificity profiles. Analysis of the 3D-structure of the DAAO enzyme from the yeast Trigonopsis variabilis (TvDAAO) revealed the Phe258 residue located at the surface of the protein globule to be in the entrance to the active site. The Phe258 residue was mutated to Ala, Ser, and Tyr residues. The mutant TvDAAOs with amino acid substitutions Phe258Ala, Phe258Ser, and Phe258Tyr were purified to homogeneity and their thermal stability and substrate specificity were studied. These substitutions resulted in either slight stabilization (Phe258Tyr) or destabilization (Phe258Ser) of the enzyme. The change in half-inactivation periods was less than twofold. However, these substitutions caused dramatic changes in substrate specificity. Increasing the side chain size with the Phe258Tyr substitution decreased the kinetic parameters with all the D-amino acids studied. For the two other substitutions, the substrate specificity profiles narrowed. The catalytic efficiency increased only for D-Tyr, D-Phe, and D-Leu, and for all other D-amino acids this parameter dramatically decreased. The improvement of catalytic efficiency with D-Tyr, D-Phe, and D-Leu for TvDAAO Phe258Ala was 3.66-, 11.7-, and 1.5-fold, and for TvDAAO Phe258Ser it was 1.7-, 4.75-, and 6.61-fold, respectively.

Evolution of universal grammar.

Universal grammar specifies the mechanism of language acquisition. It determines the range of grammatical hypothesis that children entertain during language learning and the procedure they use for evaluating input sentences. How universal grammar arose is a major challenge for evolutionary biology. We present a mathematical framework for the evolutionary dynamics of grammar learning. The central result is a coherence threshold, which specifies the condition for a universal grammar to induce coherent communication within a population. We study selection of grammars within the same universal grammar and competition between different universal grammars. We calculate the condition under which natural selection favors the emergence of rule-based, generative grammars that underlie complex language.

A new genetic variant of mdx mice: study of the phenotype.

Genetic selection in a colony of mdx mice (suffering from X-chromosome-linked muscular dystrophy) resulted in generation of their new genetic variant. In this new variant, the genetic, biochemical, and histological markers of muscular dystrophy are combined with signs of oculocutaneous albinism (skin and fur depigmentation), transillumination of the iris, sharply reduced pigmentation of the retinal epithelium, and increase of the eyeball refraction). Two sensorimotor tests (negative geotaxis and wire back down hanging) detected other phenotypical characteristics of albino mdx mice carrying, in addition to the mutation in the dystrophin gene exon 23 (intrinsic of the "classical" black mdx mice), an extra mutation responsible for pigmentation disorders. Slow geotaxis, despite longer wire back down hanging capacity, was regarded as aggravation of the neurological dysfunction in albino mdx mice in comparison with black mdx mice.

Genotype by random environmental interactions gives an advantage to non-favored minor alleles.

Fixation probability, the probability that the frequency of a newly arising mutation in a population will eventually reach unity, is a fundamental quantity in evolutionary genetics. Here we use a number of models (several versions of the Moran model and the haploid Wright-Fisher model) to examine fixation probabilities for a constant size population where the fitness is a random function of both allelic state and spatial position, despite neither allele being favored on average. The concept of fitness varying with respect to both genotype and environment is important in models of cancer initiation and progression, bacterial dynamics, and drug resistance. Under our model spatial heterogeneity redefines the notion of neutrality for a newly arising mutation, as such mutations fix at a higher rate than that predicted under neutrality. The increased fixation probability appears to be due to rare alleles having an advantage. The magnitude of this effect can be large, and is an increasing function of the spatial variance and skew in fitness. The effect is largest when the fitness values of the mutants and wild types are anti-correlated across environments. We discuss results for both a spatial ring geometry of cells (such as that of a colonic crypt), a 2D lattice and a mass-action (complete graph) arrangement.

5'-bis-pyrenylated oligonucleotides displaying excimer fluorescence provide sensitive probes of RNA sequence and structure.

Oligonucleotide conjugates bearing two pyrene residues attached to 5'-phosphate through a phosphoramide bond were synthesised. Fluorescence spectra of the conjugates show a peak typical of monomer emission (lambda(max) 382 nm) and a broad emission peak with lambda(max )476 nm, which indicates the excimer formation between the two pyrene residues. Conjugation of these two pyrene residues to the 5'-phosphate of oligonucleotides does not affect the stabilities of heteroduplexes formed by conjugates with the corresponding linear strands. A monomer fluorescence of the conjugates is considerably affected by the heteroduplex formation allowing the conjugates to be used as fluorescent hybridisation probes. The 5'-bis-pyrenylated oligonucleotides have been successfully used for investigation of affinity and kinetics of antisense oligonucleotides binding to the multidrug resistance gene 1 (PGY1/MDR1) mRNA. The changes of excimer fluorescence of the conjugates occurring during hybridisation depended on the structure of the binding sites: hybridisation to heavily structured parts of RNA resulted in quenching of the excimer fluorescence, while binding to RNA regions with a loose secondary structure was accompanied by an enhancement of the excimer fluorescence. Potentially, these conjugates may be considered as fluorescent probes for RNA structure investigation.

Solid-phase synthesis of oligoribonucleotides using T4 RNA ligase and T4 polynucleotide kinase.

The concept of solid-phase synthesis of oligoribonucleotides using T4 RNA ligase and T4 polynucleotide kinase has been proposed and tested with model homo-oligoribonucleotides. The method consists of the immobilization of the first oligomer block at the 3'-terminus on a solid support followed by a chain elongation in the 5'-direction with trinucleoside diphosphates using T4 RNA-ligase and phosphorylation using polynucleotide kinase. Hydrazides of Biogel P-300, Sepharose 4B and cellulose were tested as solid supports for immobilization of initial oligomers. The properties of supports were rated on reactivities of immobilized 5'-phosphorylated oligomers as phosphate donors in the solid phase reactions, hydrodynamical properties and capacity to eliminate donor molecules spontaneously during reactions. Hydrazide of Sepharose 4B appeared to be a more suitable support because of better hydrodynamic properties and highest reactivities of immobilized donors. Saturated concentrations of RNA ligase and polynucleotide kinase and optimal time of joining reaction were determined. In a model experiment ApApA was twice attached to the immobilized hydrazide of Sepharose 4B donor (pA)6pAox. The yield of (Ap)12 was 25%.

Natural selection of the critical period for language acquisition.

The language acquisition period in humans lasts about 13 years. After puberty it becomes increasingly difficult to learn a language. We explain this phenomenon by using an evolutionary framework. We present a dynamical system describing competition between language acquisition devices, which differ in the length of the learning period. There are two selective forces that play a role in determining the critical learning period: (i) having a longer learning period increases the accuracy of language acquisition; (ii) learning is associated with certain costs that affect fitness. As a result, there exists a limited learning period which is evolutionarily stable. This result is obtained analytically by means of a Nash equilibrium analysis of language acquisition devices. Interestingly, the evolutionarily stable learning period does not maximize the average fitness of the population.

Silver nucleolar organizer region assessment in tumor-cells from breast-cancer patients treated by extremely-low-frequency magnetic-field.

This study assessed the value of silver staining of nucleolar organizer regions (AgNORs) in tumor cells as a potential technique for the estimation of the action of ELF magnetic field on breast cancer. The light and electron microscopy was used for analysis of tissue specimens from 4 breast cancer patients after ELF magnetotherapy and 18 untreated persons with malignant and benign breast lesions. All patients had surgical removal of neoplasms. The mean number of AgNORs and the distribution of cells according to the AgNOR number in tissue samples from patients after successful ELF magnetotherapy were closer to those of persons with benign tumors, as compared to malignant ones. The effect of ELF magnetic field at the electron microscopic level is discussed.

Determination of amino acids in fodders and raw materials using capillary zone electrophoresis.

Two schemes were offered for analysis of amino acid contents in fodders and raw materials for mixed fodders by capillary zone electrophoresis (CZE). The first variant provides express analysis of four technologically important amino acids (lysine, methionine, threonine, cystine) in borate buffer on characteristic absorption of aminogroup (190 nm), with limits of quantitation being on average 0.2%. The second scheme includes pre-capillary derivatization of amino acids using phenylisothiocyanate (PITC) and separation of phenylthiocarbamyl (PTC)-derivatives obtained by CZE with a detection on 254 nm, which allows to widen a list of detectable components up to 19 (without tryptophan) and significantly improve detection limits down to 0.01%. Acid hydrolysis was used for a sample preparation. The results of analysis of fodders were compared using such methods, as CZE, ion exchange chromatography (amino acid analyzer) and reversed-phase (RP)-HPLC (with gradient technique of elution).

Long-lived reactive intermediate photogenerated from N-(5-azido-2-nitrobenzoyl)-N'-(D-biotinyl)-1,2-diaminoethane as an affinity reagent to streptavidin.

Irradiation of a complex between N-(5-azido-2-nitrobenzoyl)-N'-(D-biotinyl)-1,2-diaminoethane (I) and streptavidin with light of 313 nm led to the covalent attachment of the photobiotin analogue I to the protein. Streptavidin could also be labelled in the dark with prephotolyzed I. These results indicate that a long-lived reactive intermediate was formed upon irradiation. Moreover, after cleavage of labelled streptavidin with proteinase K this intermediate appears to be covalently attached to the same peptide as the one obtained by direct photoaffinity labelling. An iminosulfurane II derived from the reaction of biotin sulfur atom with aryl nitrene is responsible for the dark-labelling reaction. The photoproduct II converts in an aqueous solution almost completely into N-(5-amino-2-nitrobenzoyl)-N'-(D-(S-oxo)biotinyl)-1,2-diaminoethane (the half-life of II is 10 days).

5'-bis-pyrenylated oligonucleotides display enhanced excimer fluorescence upon hybridization with DNA and RNA.

A simple one-step procedure was applied for synthesis of oligonucleotide conjugates bearing two pyrene residues at the 5'-phosphate of oligonucleotide. Excimer fluorescence intensity of the conjugates is highly sensitive to duplex formation: binding of the bis-pyrenylated oligonucleotides to their DNA and RNA targets leads 10-fold increase of fluorescence. The data show that excimer fluorescence intensity of the conjugates depends linearly on the concentration of target DNA and permits quantification of DNA in solution.

PAcM-AN: poly (N-acryloylmorpholine)-conjugated antisense oligonucleotides.

A new amphiphilic, high-molecular weight poly (N-acryloylmorpholine) (PAcM) polymer has been used to be linked to oligonucleotide chains through a liquid-phase stepwise synthesis. This new conjugate has been investigated for its melting property, nuclease stability and capacity to elicit RNase H activity. Its antisense activity against an HIV-1 target has been also evaluated.

[Interaction of Escherichia coli RNA polymerase with oligoribonucleotides, homologous to "10"- and "35"- segments of the SPC promotor of bacterial genes]. / Vzaimodeistvie RNK-polimerazy Escherichia coli s oligoribonukleotidami, gomologichnymi "10;- i "35"- uchastkam SPC-promotora bakterial'nykh genov.

It was shown previously that E. coli RNA polymerase in a highly selective manner recognizes and binds 11-14-mere oligodeoxyribonucleotides related to the "-10" region of the nontranscribed DNA strand of bacterial gene promoters. The oligodeoxyribonucleotides cover the Pribnow box with flanking nucleotides up to the transcription start. These affinity oligodeoxyribonucleotides inhibit competitively the transcription of bacterial DNA carried out by E. coli RNA polymerase. The present work has demonstrated that E. coli RNA polymerase is not capable of binding the oligoribonucleotides homologous to the affinity oligodeoxyribonucleotides related to the "-10" area of the spc promoter, but binds the oligoribonucleotides which are complementary to the latter. The oligoribonucleotides with a high affinity for the E. coli RNA polymerase strongly inhibit transcription of the bacterial DNA. Attachment of alkylating groups to the 5'-ends of the affinity oligodeoxy- and oligoribonucleotides provides their covalent binding to the E. coli RNA polymerase subunits. It was shown that the modified affinity 32P-labelled oligodeoxyribonucleotide is covalently bound to the sigma-subunit while the modified affinity 32P-labelled oligoribonucleotide is covalently bound to the beta'beta-subunits of the E. coli RNA polymerase. It is suggested that the affinity oligoribonucleotides can be transcribed from the non-transcribed DNA strand in the region of the open complex and functions presumably as a primer which is splitted later from the nascent RNA or as a regulator of transcription.

[Porphyrin derivatives of oligonucleotides. I. Synthesis of oligonucleotide derivatives bearing 2,4-di(alpha-(2-hydroxyethoxy)ethyl)- deuteroporphyrin IX or a metal complex of it and study of the oxidative modification of DNA by these derivatives]. / Porfirinovye proizvodnye oligonukleotidov. I. Sintez proizvodnykh oligonukleotidov nesushchikh 2,4-di(alpha-(2-gidroksiétoksi)étil)- deiteroporfirin IX ili ego metallokompleksy i issledovanie okislitel'noi modifikatsii DKN étimi proizvodnymi.

A method for coupling 2,4-di[alpha-(2-hydroxyethoxy)ethyl] deuteroporphyrin dimethyl ether (IX), DDPOH and its complexes with metals to the 5'- or 3'-end of oligonucleotides was elaborated. In the presence of an oxidizing agent (H2O2), Fe(III)DDP-derivatives of oligonucleotides modified single-stranded DNA. The reaction was strictly site-specific and occurred at two neighbouring guanosine residues. A few types of modification were observed: cross-linking, modification leading to DNA cleavage upon piperidine treatment, and direct chain scission. The total modification yield reached 90%. Covalent attachment of Fe(III)DDP-group to oligonucleotides increased the efficiency of their uptake and the melting temperature of their complementary complexes.

[Effective complementarily-addressed photomodification of nucleic acids by oligonucleotide derivatives, containing aromatic azido groups]. / Effektivnaia komplemntarno adresovannaia fotomodifikatsiia nukleinovykhkislot proizvodnymi oligonukleotidov, soderzhashchikh aromaticheskuiu azidogruppu.

Highly effective site-specific photomodification of a DNA-target was carried out with oligonucleotide reagents carrying aromatic azido groups. Oligonucleotide derivatives with a photoactive function R on the 5'-terminal phosphate and at C-5 atom of deoxyuridine were synthesized: R1NH(CH2)3NHpd(TCCACTT) and d(ULNHRCCACTT), where R1 is p-azidotetrafluorobenzoyl, R2 is 2-nitro, 5-azidobenzoyl, R3 is p-azidobenzoyl; LNH = -CH2NH-, -CH2OCH2CH2NH- or -CH2NHCOCH2CH2NH-. The prepared compounds form stable complementary complexes and effect site-specific photomodification of the target DNA. The modification of pentadecanucleotide d(TAAGTGGAGTTTGGC) with the reagents was investigated. Maximum extent of modification strongly depended on the reagent's type, the photoreagent with R1 being the most effective. Whatever the binding site was, this agent provided a 65-70% modification in all cases except LNH = -CH2NH-, when the yield was twice lower. For the reagents bearing R1 the modification sites were identified. Selective modification at the G9 residue was detected in the case of LNH = -CH2OCH2CH2NH- and when a photoactive group was linked to the terminal phosphate.

[Diastereomers of nonionic oligonucleotide analogs. VI. Isolation of diastereomers of ethyl phosphotriesters of the octanucleotide d(GCCAAACA) by high performance affinity chromatography]. / Issedovanie diastereomerov neionnykh analogov oligonukleotidov. VI. Razdelenie diastereomerov étilovykh fosfotriéfirov oktanukleotida d(GCCAAACA) metodom vysokoéffektivnoi affinnoi khromatografii.

Diastereomers of oligonucleotide ethyl phosphotriesters were separated by high-performance complementary (affinity) chromatography on a column with the immobilized complementary oligonucleotide. The elution buffer contained 0.18 M K2HPO4, pH 7.5, and 30% acetonitrile. The temperature of the separation was a few degrees lower than Tm of corresponding oligonucleotide complexes. The diastereomers separated completely or partially were: d[GCC(Et)AAACA], d[GCCA(Et)AACA], d[GCAA(Et)ACA], d[GCC(Et)A(Et)AACA], d[GCC(Et)AA(Et)ACA], d[GCCA(Et)A(Et)ACA], d[GCC(Et)A(Et)A(Et)ACA].

[Glycogen concentration in DNA-synthesizing and non-DNA-synthesizing hepatocytes of week-old rats]. / Soderzhanie glikogena v sinteziruiushchikh i ne sinteziruiushchikh DNK gepatotsitakh nedel'nykh krysiat.

Using the combined cytochemistry, that allows to determine glycogen and DNA contents, and to localize 3H-thymidine label in one and the same cell, glycogen content was measured in both 3H-TdR-marked and 3H-TdR-non-marked hepatocytes of week-old rats. The vast majority of hepatocytes in a liver population of such rats are mononuclear 3H-TdR-non-marked cells with a 2C DNA content, which corresponds to the diploid chromosome set. The ratio of binuclear hepatocytes with diploid nuclei is in average 0.6%. At one injection of 3H-TdR, 6.6% of hepatocytes are marked in average which are almost exclusively mononuclear cells with 2c to 4c DNA content. In binuclear hepatocytes with two diploid nuclei (2c X 2), the label was seen only in single instances. The average glycogen content in S-phase hepatocytes is aproximately one third of that in hepatocytes that did not start DNA synthesis (G1 and G2 phases). The glycogen content in hepatocytes is seen reducing during S-phase. The glycogen content in G2-phase hepatocytes does not differ from that of hepatocytes before they start DNA synthesis. The availability of glycogen is not necessary for the beginning of S-phase: DNA synthesis in hepatocytes of week-old rats can start and proceed irrespective of the presence of glycogen in these. The glycogen content in binuclear hepatocytes with diploid nuclei that did not start DNA-synthesis is twice as much as in mononuclear hepatocytes before they started DNA synthesis.