Changes in the physicochemical characteristics of rabbit sclera upon scleral reinforcement

Biochemical analysis and differential scanning calorimetry demonstrated that the connective tissue (capsule) formed around a reinforcing scleroplastic implant is similar to intact sclera, its main component being type I collagen organized in perfect fibrils with cross-linking sufficient for normal thermomechanical properties. DSC also revealed a fraction of collagen with heat-labile ‘immature’ cross-links around implants containing a stimulatory plant product Panaxel, which suggested high synthetic activity of fibroblasts.     

Effect of magnesium ions on the inhibition of the mitochondrial ATPase (ATP-synthetase) complex by azide

The effect of azide on the activity and phosphorylation of ATPase in rat liver mitochondria was studied. It was shown that in the absence of exogenous Mg2+ azide inhibits both the activity and phosphorylation of ATPase. In the presence of exogenous Mg2+ azide inhibits ATPase hydrolysis, but does not inhibit the enzyme phosphorylation. It is concluded that the one-sided effect of azide on the activity of the ATPase (ATP-synthetase) mitochondrial complex is realized by participation of Mg2+. It was found also that the inhibitory effect of azide depends on the exogenous Mg2+ concentration.     

Carbonic anhydrase in leaves during radish plant ontogeny and glucose effect

Radish (Raphanus sativus L.) plants were grown in a greenhouse in soil at natural illumination, temperature, and CO₂ concentration. A relationship between the activity of soluble carbonic anhydrase (sCA) and the content of soluble carbohydrates (sCH) was investigated depending on leaf position on the stem during plant development. Indices characterizing leaf ageing during the phase of vegetative growth (PVG) (in lower leaves as compared to upper ones) were a decrease in the rate of photosynthetic release of O₂, photosynthetic efficiency, quantum efficiency of photosystem II, and the content of chlorophylls a and b. During PVG and the phase of flower bud formation, the activity of sCA and the contents of sCH, soluble protein, and Glu were the greatest in young (topmost) leaves and declined in the leaves of middle and lower storeys. From the top to the middle leaves, a dependence of sCA activity on the content of Glu or other sCH (without Glu) was direct, whereas, from the middle and the lower leaves, it became inversed. During leaf ageing in vegetatively growing plants and in the phase of budding, the relation between sCA and a relative content of Glu (percent of the rest sCH) was similar to that described above and yielded broken lines, and, during flowering, an inverse relation was observed in the leaves of all three storeys with a low content content of Glu and a low sCA activity. In order to study a dependence of sCA activity on the levels of sCH and Glu during PVG, one half of the leaf was incubated in a thin layer of water at a light schedule: by 12 h of night/day/night + 3-h-long illumination (control material). Within the period of active growth of the hypocotyl up to the storage root formation, the sCA activity and the content of sCH in the leaves incubated in water (control) increased. During a later PVG, both characteristics declined with sCA activity reduced stronger. The rate of the photosynthetic O₂ release changed in accordance with the changes in sCA activity. Incubation of the other half of the same leaf in 20 mM Glu during early PVG brought about a considerable increase in sCA activity as compared with control material. During a later PVG and at a much greater Glu concentration (330 mM), its content in the leaves increased manifold; however, there were no significant changes in the activity of sCA, the content of protein or chlorophyll. It was concluded that the activity of sCA in the radish leaves during PVG was not regulated by the absolute content of sCH or Glu in the leaves but depended on endogenous factors associated with plant development.     

Effect of surplus glucose on physiological and biochemical characteristics of sugar beet leaves in relation to the age of the leaf and the whole plant

Effect of surplus glucose on physiological and biochemical parameters of leaves of different age was investigated in sugar beet (Beta vulgaris L., subsp. saccharifera) plants in the stages of vegetative growth (SVG). Early and late SVG were differentiated by the ratio between the weights of roots and aboveground organs (0.10 and 0.35, respectively). The excess of Glu was produced by incubation of the disks excised from detached leaves in water or 0.1 M Glu at radiant flux density of 250 μmol/(m² s) with the light regime pattern described as night/day/night/light (8/16/8/3 h). In all the leaf disks incubated in water and glucose solution, the content of Glu and other soluble carbohydrates considerably increased as compared with their content in the leaves they were taken from. After disk incubation in water and glucose solution, the content of chlorophyll (a + b) rose as compared with its level in respective leaves in early SVG; in late SVG, it declined. In early SVG, the rate of the O₂ photosynthetic evolution (Ph) in the ageing leaves under saturating concentration of NaHCO₃ after incubation in water and Glu solution declined more considerably than in young leaves. In late SVG, incubation of leaf disks in water and Glu solution weakly affected P _n. The rate of O₂ dark consumption in the leaf disks of all the types of treatment increased after incubation in water and especially in Glu solution. Activity of soluble carbonic anhydrase (sCA) in the extracts from young leaves in early SVG after their incubation in water and Glu solution was essentially the same, but after the incubation of ageing leaves in Glu solution, it reliably decreased. In late SVG, sCA activity sharply decreased after incubation in water and Glu solution irrespective of the leaf age. In late SVG, activity of Rubisco in the young leaves did not change after their incubation in water but decreased after incubation of the leaves of the three ages in Glu solution. In early SVG, nonphotochemical fluorescence quenching (NPQ) in the young intact leaf was lower than in the ageing leaf, and after leaf incubation in water and Glu solution, it rose. In late SVG, the value of NPQ was greater than in early SVG and, in contrast to the leaves of early SVG, it declined after leaf incubation; in water, this decline was more pronounced than in the Glu solution. In early SVG, efficient quantum yield of photosystem II (PSII) was much greater than in late SVG and it declined in the leaves incubated with Glu. It was concluded that surplus Glu can maintain biosynthetic processes in the young leaves of young sugar beet plants (trophic function). A decline in the level of chlorophyll and the activities of sCA and Rubisco in the course of leaf development and senescence is considered as a symptom of the suppression of biosynthesis of proteins of chlorophyll-protein complexes and the enzymes (Rubisco and sCA).     

Biochemical studies on the production of neuraminidase by environmental isolates of Vibrio cholerae non-O1 from Bulgaria

Neuraminidase is a key factor in the infectious process of many viruses and pathogenic bacteria. The neuraminidase enzyme secreted by the etiological agent of cholera?- Vibrio cholerae О1?- is well studied in contrast with the one produced by non-O1/non-O139 V.?cholerae. Environmental non-O1/non-O139 V. cholerae isolates from Bulgaria were screened for production of neuraminidase. The presence of the neuraminidase gene nanH was detected in 18.5% of the strains. Тhe strain showing highest activity (30 U/mL),?V. cholerae non-O1/13, was used to investigate the enzyme production in several media and at different aeration conditions. The highest production of extracellular neuraminidase was observed under microaerophilic conditions, which is possibly related to its role in the infection of intestine epithelium, where the oxygen content is low. On the other hand, this is another advantage of the microbe in such microaerophilic environments as sediments and lake mud. The highest production of intracellular neuraminidase was observed at anaerobic conditions. The ratio of extracellular to intracellular neuraminidase production in V. cholerae was investigated. The temperature optimum of the enzyme was determined to be 50?°C and the pH optimum to be?5.6-5.8.     

The Paragon Algorithm, a next generation search engine that uses sequence temperature values and feature probabilities to identify peptides from tandem mass spectra

The Paragon Algorithm, a novel database search engine for the identification of peptides from tandem mass spectrometry data, is presented. Sequence Temperature Values are computed using a sequence tag algorithm, allowing the degree of implication by an MS/MS spectrum of each region of a database to be determined on a continuum. Counter to conventional approaches, features such as modifications, substitutions, and cleavage events are modeled with probabilities rather than by discrete user-controlled settings to consider or not consider a feature. The use of feature probabilities in conjunction with Sequence Temperature Values allows for a very large increase in the effective search space with only a very small increase in the actual number of hypotheses that must be scored. The algorithm has a new kind of user interface that removes the user expertise requirement, presenting control settings in the language of the laboratory that are translated to optimal algorithmic settings. To validate this new algorithm, a comparison with Mascot is presented for a series of analogous searches to explore the relative impact of increasing search space probed with Mascot by relaxing the tryptic digestion conformance requirements from trypsin to semitrypsin to no enzyme and with the Paragon Algorithm using its Rapid mode and Thorough mode with and without tryptic specificity. Although they performed similarly for small search space, dramatic differences were observed in large search space. With the Paragon Algorithm, hundreds of biological and artifact modifications, all possible substitutions, and all levels of conformance to the expected digestion pattern can be searched in a single search step, yet the typical cost in search time is only 2-5 times that of conventional small search space. Despite this large increase in effective search space, there is no drastic loss of discrimination that typically accompanies the exploration of large search space.     

Cyclic structural changes of endoplasmic reticulum and Golgi complex in hippocampal neurons of ground squirrels during hibernation

Repetitive remodeling and renewal of the cytoplasmic structures realizing protein synthesis accompanies the cycling of the ground squirrels between torpor and arousal states during hibernation season. Previously, we have shown the partial loss of ribosomes and inactivation of the nucleolus in pyramidal neurons in the hippocampus CA3 area at each bout of torpor with their rapid and full recovery after warming up. In the present paper, we describe reversible structural changes of the endoplasmic reticulum (ER) and Golgi complex (GC) in these neurons. The transformation of the ER form from mainly granular stacks of flattened cisternae to smooth tubules occurs at every entrance in torpor, while the reverse change happens at arousal. The torpor state is also associated with GC fragmentation and loss of their flattened cisternae, i.e., dictiosomes. In neurons, the appearance of the autophagosomal vacuoles containing fragments of membrane structures and ribosomes in torpor state is a sign of the partial destruction of ER and GC. Granular ER restoration, perhaps through assembly from the multilamellar membrane structures, bags or whorls begins in the middle of the torpor bout, while GC dictiosomes reappear only during warming. The ER and GC completely restore their structure 2–3 h after the beginning of arousal. Thus, hibernation represents an example of the structural adaptation of the nerve cell to deep changes in functional and metabolic activity through both the active destruction and renewal of ribosomes, ER, and GC. Perhaps, namely the incomplete ER autophagosomal degradation in torpor provides its rapid renewal at arousal through the reassembly from the preserved fragments.     

Selenoprotein W depletion induces a p53- and p21-dependent delay in cell cycle progression in RWPE-1 prostate epithelial cells

The anticancer activity of selenium (Se) has been demonstrated in myriad animal and in vitro studies, yet the mechanisms remain obscure. The main form of Se in animal tissues is selenocysteine in selenoproteins, but the relative importance of selenoproteins versus smaller Se compounds in cancer protection is unresolved. Selenoprotein W (SEPW1) is a highly conserved protein ubiquitously expressed in animals, bacteria, and archaea. SEPW1 depletion causes a delay in cell cycle progression at the G1/S transition of the cell cycle in breast and prostate epithelial cells. Tumor suppressor protein p53 is a master regulator of cell cycle progression and is the most frequently mutated gene in human cancers. p53 was increased in SEPW1 silenced cells and was inversely correlated with SEPW1 mRNA in cell lines with altered SEPW1 expression. Silencing SEPW1 decreased ubiquitination of p53 and increased p53 half-life. SEPW1 silencing increased p21(Cip1/WAF1/CDKN1A), while p27 (Kip1/CDKN1B) levels were unaffected. G1-phase arrest from SEPW1 knockdown was abolished by silencing p53 or p21. Cell cycle arrest from SEPW1 silencing was not associated with activation of ATM or phosphorylation of Ser-15 in p53, suggesting the DNA damage response pathway was not involved. Silencing GPX1 had no effect on cell cycle, suggesting that G1-phase arrest from SEPW1 silencing was not due to loss of antioxidant protection. More research is required to identify the function of SEPW1 and how it affects stability of p53.     

Sexual and reproductive behaviour of Drosophila melanogaster from a microclimatically interslope differentiated population of "Evolution Canyon" (Mount Carmel, Israel).

The strong microscale interslope environmental differences in "Evolution Canyon" provide an excellent natural model for sympatric speciation. Our previous studies revealed significant slope-specific differences for a fitness complex of Drosophila. This complex involved either adaptation traits (tolerance to high temperature, different viability and longevity pattern) or behavioural differentiation, manifested in habitat choice and non-random mating. This remarkable differentiation has evolved despite a very small interslope distance (a few hundred metres only). Our hypothesis is that strong interslope microclimatic contrast caused differential selection for fitness-related traits accompanied by behavioural differentiation and reinforced by some sexual isolation, which started incipient speciation. Here we describe the results of a systematic analysis of sexual behaviour in a non-choice situation and several reproductive parameters of D. melanogaster populations from the opposite slopes of "Evolution Canyon". The evidence indicates that: (i) mate choice derives from differences in mating propensity and discrimination; (ii) females from the milder north-facing slope discriminate strongly against males of the opposite slope; (iii) both sexes of the south-facing slope display distinct reproductive and behavioural patterns with females showing increased fecundity, shorter time before remating and relatively higher receptivity, and males showing higher mating propensity. These patterns represent adaptive life strategies contributing to higher fitness.     

Purification and characterization of a novel sialidase from a strain of Arthrobacter nicotianae

The nonpathogenic strain Arthrobacter nicotianae produces two sialidase isoenzymes, NA1 and NA2, with molecular masses of 65 kDa and 54 kDa, respectively, as determined by 10% SDS-polyacrylamide gel electrophoresis. NA1 and NA2 exhibit maximum activities at pH 4 and 5, and both show clear thermal optima at 40 degrees C. They are stable at temperatures up to 50 degrees C. The critical temperatures (T (c) = 50 degrees C and 51 degrees C) for the two isoenzymes were determined by fluorescence spectroscopy and correlate well with the temperatures of melting (T (m) = 49 degrees C and 48 degrees C), determined by CD spectroscopy. The isoenzymes are less stable against denaturation with Gdn.HCl, and the free energy of stabilization in water was calculated to be 7.6 and 8.0 kJ mol(-1), respectively. The specific activity (K (m) value) toward glucomacropeptide as a substrate was calculated to be 0.126 mM for NA1 and 0.083 mM for NA2.