Comparative studies on thymidylate synthetase from P. berghei and mouse reticulocytes

Partially purified thymidylate synthetase from Plasmodium berghei and mouse reticulocytes was characterized. The mol. wt of the enzyme from P. berghei was about twice that from mouse reticulocytes. The optimum pH of the enzyme from P. berghei was found to be 6.5-7.5 while that from the host was 7.0-8.0. The enzyme from P. berghei was more susceptible to pH denaturation than the enzyme from reticulocytes. The enzyme from both sources differed in their Km values for substrates. The enzyme from reticulocytes was less sensitive to inhibition by substrate analogs than that from P. berghei.     

江淮丘陵地区油-稻轮作条件下土壤硫库变化研究

江淮丘陵地区下蜀系黄土母质发育的水稻土油2稻轮作试验表明, 油菜种植期间, 耕层土壤硫主要来源于耕层以下土层的补给, 其次是大气干湿沉降;该时期耕层土壤硫输出主要是油菜吸收, 其次是淋失.油菜种植期间耕层土壤硫输入量小于输出, 导致耕层土壤硫库下降8.76kg·hm^-2, 22%来自无机硫库的下降.水稻种植期间, 耕层土壤硫输入主要来自灌溉水, 其次是底土层的补给和大气干湿沉降;而硫输出主要是淋失, 其次是水稻吸收.耕层土壤硫输入量大于输出, 导致耕层土壤硫库增加18.69kg·hm^-2, 18%来自无机硫库的增加.全年油2稻轮作期间耕层土壤硫输入量大于输出, 导致耕层土壤硫库增加9.93kg·hm^-2, 13%来自无机硫库的增加.     

Effect of ethanol on activity of the plasma-membrane ATPase in, and accumulation of glycine by, Saccharomyces cerevisiae

The pH optimum of the ATPase activity in plasma membranes from Saccharomyces cerevisiae NCYC 431 from 8 h cultures was around 6.5 and that in membranes from organisms from 16 h cultures near 6.0. The Km[ATP] of the enzyme was virtually unaffected by the age of the culture from which organisms were harvested, although the Vmax of the enzyme in membranes from organisms from 8 h cultures was higher than that for organisms from 16 h cultures. Ethanol non-competitively inhibited ATPase activity in membranes, although the inhibition constant for the enzyme from organisms from 8 h cultures was lower than that from organisms from 16 h cultures. Glycine accumulation by the general amino acid permease was non-competitively inhibited by ethanol. Inhibition constants were virtually the same for glycine uptake by deenergized organisms from 8 h and 16 h cultures, but under energized conditions the value was greater for organisms from 16 h rather than 8 h cultures. The data indicate that inhibition of plasma-membrane ATPase activity by ethanol could account, at least in part, for inhibition of glycine accumulation by ethanol.     

精制破伤风类毒素产毒及精制工艺的改进

用超滤、硫酸铵二段盐析法取代等电点沉淀法后,精制破伤风类毒素（精破类）的纯度由８０７Ｌｆ／ｍｇＰＮ提高到１８８３Ｌｆ／ｍｇＰＮ,纯度提高一倍以上。使用双胨培养基取代酪素培养基后,产毒水平由４７Ｌｆ／ｍｌ提高到８８Ｌｆ／ｍｌ（ｔ＝６．４６,ｐ＜０．００１）;用新法精制后,精破类纯度分别为１９４９Ｌｆ／ｍｇＰＮ及１７８５Ｌｆ／ｍｇＰＮ（ｔ＝０．３３４,ｐ＞０．０５）,引用双胨培养基后可提高产毒水平,但不影响精破类的纯度。     

Variability of biological glucose transport activity in human erythrocytes]

Glucose transport activity was determined on erythrocytes from healthy adult and children. A large variability of values was found between each donor, but transport rate was significantly higher in erythrocytes from adults than those from children. In opposite, the same number of glucose transport sites was found on erythrocytes from adult and children, suggesting that transporters from adult erythrocytes exhibited an higher own activity than transporters from child erythrocytes. Involvement of various factors in glucose transport activity was discussed.     

Arginase from rat fibrosarcoma. Purification and properties

Arginase from rat fibrosarcoma was purified about 1900-fold and its properties were compared with those of the enzyme from liver and kidney. Arginase from fibrosarcoma was a neutral protein of molecular weight 120,000 with a K_m value of 11 mM for arginine. The activation energy was 7.2 kcal/mol and the pH optimum was 10. The fibrosarcoma enzyme was immunologically different from that of the liver. The arginase from fibrosarcoma closely resembled the arginase from the kidney in its electrophoretic, kinetic and immunological properties.     

Use of collagenase from the hepatopancreas of the Kamchatka crab for isolating and culturing endothelial cells of the large vessels in man

The application of hepatopancreas collagenase from crab Paralithodes camtschatica for the isolation and cultivation of the endothelial cells was studied in human umbilical vein endothelium. The comparison of the enzyme from crab hepatopancreas with collagenase from Clostridium histoliticum has shown that the number of viable cells isolated from human umbilical vein by the crab enzyme was lower than in the case of microbial collagenase. However, this difference was not significant for subsequent cultivation of cells. Harvesting of the endothelial cells from the substrate during cultivation was more effective in the case when collagenase from crab hepatopancreas was used. It was shown that crab collagenase, in contrast with microbiological collagenase, was not a metal-dependent enzyme.     

Increased phosphorylation of a specific protein in mitogen-stimulated lymphocytes

The phosphorylation of proteins from murine splenic lymphocytes was studied. When the phosphorylation of proteins in extracts from Concanavalin A (ConA)-treated lymphocytes was compared with that of resting lymphocytes, there was only one detectable difference between them. A protein of 135,000 mol. wt was highly phosphorylatable in extracts from ConA-treated cells while phosphate incorporation into this protein was slight in extracts from untreated cells. This effect could be observed 12 h after ConA treatment and was maximal during S phase. This soluble protein was also phosphorylated in intact lymphocytes.     

Comparison of C-S lyase in Lentinus edodes and Allium sativum

The characteristics of C-S lyase in Lentinus edodes (shiitake) were compared with those in Allium sativum (garlic). C-S lyase mRNA from shiitake was hybridized with the garlic C-S lyase cDNA fragment, being almost the same length as that from garlic. The isoelectric point of the C-S lyase from shiitake was between pH 4 and 5, while that from garlic was over a wider range between pH 4 and 8. Different from the C-S lyase from garlic, that from shiitake was not a glycoprotein without being stained by PAS, and was not bound to the anti-garlic C-S lyase antibody. Similar to garlic C-S lyase, shiitake C-S lyase comprised a homodimer, and its molecular mass was 84 kDa. However, the N-terminal amino acid sequences of each subunit of shiitake C-S lyase were totally different from those of garlic C-S lyase.     

Mutarotation of hydrolysis products by different types of exo-cellulases from Trichoderma viride.

Mutarotation of products from p-nitrophenyl beta-D-cellobioside and cellopentaitol by two different types of exo-cellulases from Trichoderma viride was investigated. It was found that an exo-cellulase of glucosidase type produced from the former substrate D-glucose which was mutarotated in a downward direction, while another exo-cellulase of Avicelase type produced from the latter substrate cellobiose which was mutarotated in an upward direction.     

Genetic variation in Beauveria bassiana populations associated with the darkling beetle, Alphitobius diaperinus

A study was conducted to assess genetic variation within and among populations of Beauveria bassiana (Deuteromycotina: Hyphomycetes) associated with the darkling beetle, Alphitobius diaperinus (Coleoptera: Tenebrionidae), using RAPD markers. A hierarchical collection of samples (strains from the same insect specimen, from insects from the same location, and from insects from different locations) was obtained from infected beetles from North Carolina (NC) and West Virginia (WV), USA. Ten primers resolved 81 strains into 80 distinct multiband phenotypes reflecting the substantial amount of variation that was present. Variation present within populations was evident not only in the separation of each strain as a distinct multiband phenotype but also in the separation of strains within a population into separate clusters. Among populations, a group sharing more than 89% similarity was observed among all the strains from Martin Co. and Greene Co., NC and 61% of the strains collected from WV. Some genetic differentiation was present among the other populations but the separation was not distinct with a few strains from some populations showing greater affinity to strains from other collection sites.     

Ferredoxin-dependent methane formation from acetate in cell extracts of Methanosarcina barkeri (strain MS).

Cell extracts of Methanosarcina barkeri grown on acetate catalyzed the conversion of acetyl-CoA to CO2 and CH4 at a specific rate of 50 nmol min-1 mg-1. When ferredoxin was removed from the extracts by DEAE-Sephacel anion exchange chromatography, the extracts were inactive but full activity was restored upon addition of purified ferredoxin from M. barkeri or from Clostridium pasteurianum. The apparent Km for ferredoxin from M. barkeri was determined to be 2.5 M. A ferredoxin dependence was also found for the formation of CO2, H2 and methylcoenzyme M from acetyl-CoA, when methane formation was inhibited by bromoethanesulfonate. Reduction of methyl-coenzyme M with H2 did not require ferredoxin. These and other data indicate that ferredoxin is involved as electron carrier in methanogenesis from acetate. Methanogenesis from acetyl-CoA in cell extracts was not dependent on the membrane fraction, which contains the cytochromes.     

Colour remediation of pulp mill effluent using purified fungal cellobiose dehydrogenase: reaction optimisation and mechanism of degradation

Cellobiose dehydrogenase purified from two different fungal sources was assessed for its ability to remove and/or reduce colour from pulp mill bleach plant effluent. Cellobiose dehydrogenase purified from Phanerochaete chrysosporium was shown to prefer acidic conditions and was consequently used to treat the acid effluent stream discharged from a pulp mill bleach plant, while an analogous enzyme originating from Humicola insolens preferred alkaline conditions, and was applied to the effluent discharged from the caustic sewer of the bleach plant. Both enzyme preparations were able to remove colour from their respective effluent sources to a comparable extent. Up to 50% of the effluent colour was removed within 4 days when treated under optimised conditions. Furthermore, it was also shown that this enzymatic approach was effective at removing colour generated by both softwood and hardwood resources. Mechanistically, it was shown that colour was removed from all molecular weight fractions, and the higher molecular weight material (>300 kDa) was concurrently preferentially degraded. Cellobiose dehydrogenase treatment of effluent did not target phenolic, stilbene, or alpha-carbonyl structures, but did affect the quinone content. Further investigations using model compounds confirmed these results, and subsequently showed that only the para-quinones with low substitution were reduced with cellobiose dehydrogenase.     

Ca/Mg-dependent endonuclease as a probe for detecting chromatin changes in lymphocytes in chronic lymphoid leukemia

Chromatin fragmentation of bovine peripheral blood lymphocytes from normal animals and the ones suffering from chronic lympholeucosis (CLL) by DNase I, micrococcal nuclease and purified Ca/Mg-dependent endonuclease from nuclei of human splenocytes was studied. The lymphocytes chromatin from CLL animals was shown to be more resistant to nucleases, than the one from normal animals. It was found that difference between fragmentation of chromatin samples from normal and CLL bovines was more dramatic when Ca/Mg- dependent endonuclease was used versus traditionally exploited DNase I and micrococcal nuclease. The data suggest that purified Ca/Mg-dependent endonuclease can be a useful enzymatic probe for detection of lymphocytes chromatin changes during CLL.     

Mineralization of copper, manganese and zinc from rock mineral flour and city waste compost for efficient use in organic farming

Restricted supply of micronutrients is a common constraint for plant growth worldwide, especially in organic farming systems where nutrients supply to crops mostly depends on the mineralization of native soil organic matter, decomposition of applied manures and crop residues. A laboratory incubation study was therefore conducted to investigate the potential release of copper (Cu), manganese (Mn) and zinc (Zn) from the rock mineral flour (RMF) and city waste compost (CWC) as compared to inorganic micronutrient fertilizers for 140 days. Release of the micronutrients from RMF and CWC showed different trends. The results showed that about 4.6% of Cu added as RMF was released irrespective of the quantity of the RMF applied. However, Cu release from CWC increased from 0.7 to 3.5% as the amount of compost added was increased. Copper recovery from copper sulphate was 98%. Manganese release from RMF decreased from 114 to 103% as the RMF level was increased, while the corresponding decrease in Mn release from CWC was from 14 to ?3%. Manganese recovery from manganese sulphate was 100%. Zinc release from RMF increased from 5.8?15.5%, with an increase in the amount of RMF applied, while no Zn was released from CWC. Recovery of Zn from zinc sulphate was 98%. These results show that RMF and CWC could be used to meet Cu, Mn and Zn requirements of organically grown cereals. The results of the investigation have general applicability in organic farming.     

Differences between HMG1 proteins isolated from normal and tumour cells

The properties of the non-histone chromosomal high-mobility-group 1 (HMG1) proteins from rat liver and Guerin ascites tumour cells (GAT cells) were compared and showed the following differences: (1) five spots were missing in the peptide map of HMG1 from GAT cells in comparison with that of HMG1 from rat liver; (2) HMG1 from GAT cells was about 5-times more poly(ADP)-ribosylated; (3) HMG1 from GAT cells which was found acetylated in vivo and incorporated [14C]acetate in vitro, whereas no incorporation of the label was detected in HMG1 from rat liver; (4) HMG1 from GAT cells exhibited pronounced ability to form oligomers at physiological ionic strength, while HMG1 from rat liver was predominantly in monomeric form. This property of HMG1 from GAT cells was lost upon deacetylation.     

Chemotactic function of skin fibroblasts in patients with amyloidosis

Chemotaxis of cultivated fibroblasts, obtained from patients with amyloidosis, chronic glomerulonephritis and healthy volunteers, was investigated. Fibroblast migration toward donor serum and serum from patients with amyloidosis was measured using Boyden chamber's technique. As "zero" chemoattractant Hank's solution was used. It was shown, that chemotactic index (CI) was independent from cell density. Significant CI depression of fibroblasts from patients with amyloidosis toward donor serum in contrast to fibroblasts from patients with chronic glomerulonephritis and healthy volunteers was shown. The depression of chemotactic function was the same with fibroblasts from patients with different variants of amyloidosis and different stages of amyloid nephropathy and was stable in several cell generations. The results obtained suggest the existence of primary hereditary variant (variants) of chemotactic function, which may lead to the development of amyloidosis in certain conditions.     

Phosphoenolpyruvate Carboxylase from Mesembryanthemum crystallinum: Its Isolation and Inactivation in vitro

Phosphoenolpyruvate carboxylase (PEPC) was isolated from leavesof Mesembryanthemum crystallinum, which performed Crassulaceanacid metabolism. PEPC was much more stable when extracted from expanded thanfrom expanding leaves. The inactivation of PEPC in desaltedextracts from expanding leaves was much faster at 25 than at0 °C, was stimulated by raising Mg²⁺ from 0.1 to 3.0 mM,and was reduced by bovine serum albumin. The same type of inactivationwas found after mixing extracts from the two types of leaves,and the decrease in PEPC activity then also included inactivationof the ’stable’ PEPC from the expanded leaves. Afterelution from DEAE-cellulose, PEPC from expanding leaves wasmuch more stable than in desalted, crude extracts. It is suggested that another enzyme is involved in this inactivationof PEPC, but this needs verification.