183 Isolation and partial characterization of a general hormone transporting blood protein complex

Using immobilized insulin, human blood serum insulin-binding proteins were isolated at physiological pH values and zinc concentrations. By means of enzyme-linked immunoassay, albumin (26.7?±?1.75%), α-fetoprotein (2?±?0.61%), transferin (10?±?0.73%), retinol-binding protein (4?±?0.75%), and immunoglobulin G (20.3 ± 0.95%) were identified in isolated protein complex from human blood serum. Gel filtration of isolated proteins on Sephadex G-75 has demonstrated formation of a supramolecular complex under pH 7.2 and zinc concentration of 1?mg/mL (Garipova et al., 2010). The possible existence of such complex is supported by the presence of lipocalin glycoproteins such as retinol-binding protein and transferrin in obtained proteins; lipocalins form the protein family, which is capable to form intermolecular complexes to ensure the delivery of hormones to target cells through their own cell receptors (Flower, 1996). It has been shown that the isolated protein complex, along with insulin, transports other hydrophilic hormones. By means of ELISA, protein hormones such as luteinizing (1?mU/mg), thyreotropic (0.03?mM/mg) hormones, and prolactin (0.5?mU/mg) were identified. In addition, the complex has included hydrophobic hormones: thyroxin (11.0?±?0.5?nmol/mg), triiodothyronine (1.3?±?0.06?nmol/mg), and testosterone (7.0?±?0.03?nmol/mg). Based on these data, we have proposed an existence in human blood of a general transporting complex that is common for both hydrophobic and hydrophilic hormones. The core of this complex is formed by transporting proteins, including those relating to lipokalins. This complex transfers thyroxin, triiodothyronine, testosterone, thyroid-stimulating hormone, prolactin, and luteinizing hormone, as well as triglycerides and cholesterol to the tissues. The composition of insulin-binding protein complex was significantly changed in the insulin-dependent diabetes mellitus patient’s blood. Albumin and α-fetoprotein, which are present in a normal complex, are revealed only in trace amounts in samples isolated from the diabetic blood and are replaced by α₁-acid glycoprotein, and possibly other unidentified proteins. Disturbance of hormone-transporting complex protein composition in the diabetic blood may be a reason of insulin delivery disruption. Perhaps, for the same reason, delivery to target cells of not only insulin, but also other members of the hormone-transporting complex, is disrupted in the diabetic blood.     

Hypoxia and Imbibition Injuries to Aging Seeds

The development of hypoxia and primary injuries were examined during the imbibition of aging pea seeds (Pisum sativum L., cv. Nemchinovskii). The distribution of air-dry pea seeds by their room-temperature phosphorescence revealed the presence of two fractions (I and II) in a seed lot with 72% germinability and three fractions (I, II, and III) in a seed lot with 50% germinability. The water uptake during imbibition was slower in the fraction I seeds than in the fraction-II seeds. The fraction-I seeds produced normal seedlings, whereas the fraction-II seeds either produced seedlings with morphological defects (abnormal) or did not germinate at all. The fraction-III seeds were all dead. The phosphorescence of endogenous porphyrins, emitted only at low O₂ content, was measured after 20-h seed imbibition. The fraction-I seeds emitted no discernible phosphorescence. The fraction-II comprised highly phosphorescent seeds incapable of radicle protrusion and moderately phosphorescent seeds producing abnormal seedlings. The fraction-II seeds experienced hypoxia during the imbibition because of rapid oxygen consumption by the embryo and restrictions to O₂ diffusion imposed by the seed coat. In the fraction-I seeds, the rate of oxygen consumption by the embryo was slower and the seed coat resistance to oxygen diffusion was lower than in the fraction-II seeds. Therefore, hypoxia did not arise in the fraction-I seeds. The submergence of seeds in water caused lethal injuries. The imbibition of seeds without any contact with water caused no lethal damages but did not reduce the percentage of seeds dying of hypoxia. A slow imbibition of seeds in the media containing either an osmoticum (PEG) or an inhibitor of aquaporin channels (p-chloromercuribenzoate) prevented the lethal injuries at early stages of seed hydration and retarded the appearance of oxygen deficiency in fraction-II seeds. Different rates of water uptake by fraction-I and fraction-II seeds were controlled by permeability of cell membranes rather than by permeability of seed coat. It is proposed that low permeability of plasma membranes to water in fraction-I seeds results from the predominantly closed aquaporin channels, whereas a higher permeability of weak seeds (fraction II) is due to open channels.     

Insertion-deletion polymorphism of the angiotensin converting enzyme gene and its relationship to serum levels of free amino acids in the patients with connective tissue dysplasias

An association between insertion/deletion polymorphism (IDP) of the Alu repeat in intron 16 of the angiotensin I-converting enzyme (ACE) gene and the serum free amino acid levels in the patients with connective tissue dysplasias was examined. Genotyping of 102 patients (25 II, 51 ID, and 26 DD) was performed using PCR. Serum free amino acids levels in these patients were determined by use of HPLC technique. A statistically significant increase of the leucine-isoleucine (P < 0.05) and phenylalanine (P < 0.01) levels in deletion homozygous patients (DD) relative insertion homozygous (II) patients was observed. The differences in respect of other amino acids were not detected. These findings point to the importance of registration of IDP in the ACE gene at dietary therapy of such patients, as well as in the individual choice of medical preparations containing the amino acids mentioned.     

Variability of microsatellites BM224 and Bcal7 in populations of green toads (Bufo viridis complex) differing by nuclear DNA content and ploidy

The variability of microsatellites BM224 and Bcal7 was studied for the first time in three species of the diploid-polyploid complex of Bufo viridis (B. viridis, B. oblongus, and B. pewzowi). The locus Bcal7 was established to be monomorphic in all samples studied. In microsatellite BM224, three allele variants were found. Among tetraploid toads, the western Asiatic species B. oblongus was characterized by one allele only, the eastern B. pewzowi, by the two other alleles. A similar distribution was also revealed in triploid individuals on the borders of range between tetraploid and diploid species. Among the diploid species B. viridis samples, all three allele variants of microsatellite BM224 were observed. Their distribution in the area proved to be geographically determined. In diploid toads, a similarity was revealed between the distribution of microsatellite BM224 alleles and variability of the nuclear DNA content.     

Method and equipment for detection of pathogenic microorganisms using tracking analytical membranes

Method for detection of pathogenic microorganisms in different microbiological assays (water, faeces, punctate etc.) has been developed. Device for preparation of samples and filtration of studied material through transparent tracking membranes with fixed size of pores and subsequent microscopy of filtrate was developed also as well as device for filtration of different liquids in closed cylinder under pressure. Method significantly increases informativeness and reliability of analyses, and decreases of their labor and financial costs.     

Investigation of some impurity profiles by (n,α) reactions

To investigate some impurity profiles by (n,α) reactions for destructive and nondestructive analysis, two experimental facilities on a horizontal channel of the well-moderated research reactor have been developed. In accordance with an α-particle registration geometry two different measurement techniques based on using a surfacebarrier Si(Au) detector and a counting ionization chamber have been used. The determination limit of boron is ofn x 10¹⁶ cm^-3 or (1 /dv 5) x 10^-5 % with a depth resolution of ±0.03 Μm in semiconductors. A solid-state nuclear track detector (SSNTD) has also been used to determine the boron surface and deep distributions in semiconductors, plants, and other materials.     

Regulation of terephthalate catabolism in Rhodococcus rubropertinctus

The regulation of terephthalate catabolism was studied in Rhodococcus rubropertinctus which decomposed this synthetic monomer. The pathway (a) of terephthalate (TP) catabolism is as follows: TP----benzoate----4-hydroxybenzoate----protocatechuate----pyrocatechol-- --cycle ortho-cleavage. The following results were obtained when studying why two other catabolic pathways were realized if benzoate and 4-hydroxybenzoate were taken as a sole carbon source, namely, (b) benzoate----pyrocatechol----cycle cleavage and (c) 4-hydroxybenzoate----protocatechuate----cycle cleavage. TP seemed to cause the divergence of pathways (a) and (b) by repressing the system of benzoate oxidation to pyrocatechol. In pathway (c), benzoate repressed the synthesis of enzymes which catalysed protocatechuate oxidation. Pathway (b) was switched over to (a) when the strain was grown in a medium containing TP and benzoate at a benzoate concentration above 5 mM. Here, the concentration of benzoate (first exogenous and later formed from TP) played a key role. R. rubropertinctus growth in a medium with TP and glucose had diauxic characteristics.     

Variations in <Emphasis Type="Italic">BM224</Emphasis> microsatellite in green frogs of genus <Emphasis Type="Italic">Rana</Emphasis>

The variable microsatellite repeat BM224 has been revealed in the genomes of eight species of green frogs (Rana ridibunda, R. cf. bedriagae, R. cretensis, R. esculenta, R. lessonae, R. shquiperica, R. saharica, R. nigromaculata). Previously, this repeat was observed in members of the genus Bufo. In this paper, the possibility of using this genetic marker for species identification is discussed.