转铁蛋白受体单链抗体与BDNF融合蛋白的表达及活性鉴定

脑源性神经营养因子（BDNF）对中枢神经系统的多种神经元具有营养,修复和保护功能,但因无法通过血脑屏障限制了其应用。本文利用抗转铁蛋白受体（TfR）的单链抗体（ox26-scFv）作为脑转运载体,分别扩增单链抗体和BDNF基因,插入pTIG-Trx载体,构建融合基因表达载体pTIG-Trx/scFv-BDNF,在大肠杆菌BL21（DE3）中实现了高效表达。经Ni-NTA金属鏊合层析柱纯化后,在41Kd处可见目的纯化条带。大鼠GH3细胞免疫酶染色显示,ScFv-BDNF融合蛋白能与转铁蛋白受体特异性结合。同时能够促进鸡胚背根节神经突起的生长,具备了BDNF的生物学活性。为使BDNF能够跨越血脑屏障成为中枢神经系统的治疗药物打下了实验基础。     

维生素E对三氯乙烯急性染毒大鼠肝脏脂质过氧化与抗氧化酶的影响

本文观察了用抗氧化剂维生素E预处理后,三氯乙烯(3000mg/kg B-W-) 一次性经口染毒24h 大鼠肝脏的超氧化物歧化酶、谷胱甘肽过氧化物酶等抗氧化酶活力及丙二醛含量的变化,结果表明三氯乙烯染毒组肝脏中丙二醛含量、超氧化物歧化酶活力及血清谷丙转氨酶、谷草转氨酶活力均高于对照组(P< 0-01) ;而维生素E干预组的丙二醛含量、超氧化物歧化酶活力及血清谷丙转氨酶、谷草转氨酶活力均分别低于三氯乙烯染毒组(P< 0-01) ,说明三氯乙烯急性染毒可引起肝脏脂质过氧化反应及肝损害,肝脏超氧化物歧化酶活力升高可能是机体受自由基及脂质过氧化反应刺激而诱导产生的一种适应性反应,维生素E对三氯乙烯所致的肝损害有一定的保护作用。     

白腐菌木质素降解酶及其在木质素降解过程中的相互作用

木质素是一类不易降解的生物物质,在自然界中,白腐真菌对木质素的降解能力最强.白腐真菌降解木质素主要依靠分泌的三种酶:木质素过氧化物酶(Lip)、锰过氧化物酶(MnP)和漆酶(Lac).对白腐真菌分泌的三种木质素降解酶在性质、分布等方面进行了比较,系境地介绍三种木质素降解酶的催化作用,并阐述其在木质素降解过程中的相互作用.     

Chloride-bicarbonate exchange in the urinary bladder of the turtle. Independence from sodium ion

The rates of Cl^? absorption and HCO^?₃ secretion were not different in turtle urinary bladders bathed in Na⁺-containing and solutions.These results in turtle bladder are inconsistent with Na⁺-anion cotransport but can be accounted for by a Cl^?/HCO^?₃ exchange system.     

蒙古沙鼠脑缺血后大脑皮质Calretinin(CR)的表达变化

目的:观察蒙古沙鼠前脑短暂缺血后前额皮质Calretinin(CR)的表达变化,为进一步研究其功能及治疗提供形态学依据。方法:24只健康雄性长爪鼠随机分为正常组对照组(6只)和缺血组(18只),夹闭蒙古沙鼠双测颈总动脉10 min诱导前脑缺血后,动物分别存活1天、3天或7天。用免疫组织化学方法检测了前额皮质中CR的表达。结果:与正常组相比,各缺血组中前额皮质中CR表达均上调。缺血再灌1天时,CR表达最强(P＜0．01);3天时CR表达开始恢复(P＜0．05);7天时CR表达进一步恢复,但仍高于正常组(P＜0．01)。结论:前脑短暂缺血后可造成蒙古沙鼠前额皮质CR的表达在短时间内急剧上调;但随着时间的CR的表达会恢复正常。     

HL－60细胞诱导分化过程中蛋白质酪氨酸磷酸化水平的改变

对ＲＡ、ＨＨＴ和ＷＢ_（６５２）诱导ＨＬ－６０细胞过程中,细胞浆和膜溶脱部分的蛋白质酪氨酸磷酸化水平变化进行了对比研究,结果发现,在胞浆部分主要有四种含有Ｐ－Ｔｙｒ的蛋白,而且８０ｋＤ蛋白酪氨酸磷酸化水平随着诱导发生变化。诱导前后内源性蛋白上Ｐ－Ｔｙｒ百分含量也发生了改变。     

Synthesis and characterization of a high spin d Fe(II) complex with the tridentate ligand dipyrido(2,3-a:3,2-j)phenazine (dpop)

A new bis-(N-tridentate) Fe(II) complex [Fe(dpop^′)₂](PF₆)₂ (dpop^′=dipyrido(2,3-a:3^′,2^′-j)phenazine) was prepared and studied. The magnetic moment of the solid was determined as μ=5.2-4.9 BM and in CH₃CN solution as μ=4.9 BM and indicate the high spin Fe(II) state. The electronic absorption spectrum displays a broad weak absorption MLCT transition at 602 nm (ε=3.8×10³ M⁻¹ cm⁻¹), consistent with CT absorptions of other Fe(II) HS complexes. The cyclic voltammogram of the complex shows an irreversible Fe^2+/3+ oxidation at +1.55 V and two dpop^′0/−1 centered reductions at −0.20 and −0.59 V versus Ag/AgCl.     

Cr(VI) reduction by Enterobacter sp. DU17 isolated from the tannery waste dump site and characterization of the bacterium and the Cr(VI) reductase

Out of nineteen bacteria screened from the tannery waste dump site, the most effective isolate, strain DU17 was selected for Cr(VI) reduction process among the non-pathogenic once. Based on 16S rRNA gene sequence analysis, the bacterium was identified as Enterobacter sp. DU17. Its amplified Cr(VI) reductase gene showed maximum homology with flavoprotein of Enterobacter cloacae. Enterobacter sp. DU17 reduced Cr(VI) maximally at 37 °C and pH 7.0. Various co-metals, electron (e⁻) donors and inhibitors were tested to study their effect on Cr(VI) reduction. In presence (0.2% each) of glucose and fructose, Enterobacter sp. DU17 reduced Cr(VI) completely after 16 and 20 h, respectively. Since the concentration of total Cr was invariable after remediation as detected through AAS analysis, this experiment disclosed that responsible operation was associated with extracellular Cr(VI) reduction process rather than uptake mechanism. Multiple antibiotic resistance index of 0.08 for this bacterium was very low as compared to standard risk assessment value of 0.20. With high Cr(VI) reducing capability, non-pathogenicity and antibiotic sensitivity, Enterobacter sp. DU17 is found to be very efficient in removing Cr(VI) toxicity from the environment.     

Letting the gene out of the bottle: the population genetics of genetically modified crops

Genetically modified (GM) plants are rapidly becoming a common feature of modern agriculture. This transition to engineered crops has been driven by a variety of potential benefits, both economic and ecological. The increase in the use of GM crops has, however, been accompanied by growing concerns regarding their potential impact on the environment. Here, we focus on the escape of transgenes from cultivation via crop x wild hybridization. We begin by reviewing the literature on natural hybridization, with particular reference to gene flow between crop plants and their wild relatives. We further show that natural selection, and not the overall rate of gene flow, is the most important factor governing the spread of favorable alleles. Hence, much of this review focuses on the likely effects of transgenes once they escape. Finally, we consider strategies for transgene containment.     

The subunit interface of the Escherichia coli ribosome: Identification of proteins at the interface between the 30 S and 50 S subunits by crosslinking with 2-iminothiolane

The 70 S ribosomes of Escherichia coli were treated with 2-iminothiolane with the resultant addition of 110 sulfhydryl groups per ribosome. The modified ribosomes were oxidized to promote disulfide bond formation, some of which formed intermolecular crosslinks. About 50% of the crosslinked 70 S ribosomes did not dissociate when exposed to low concentrations of magnesium in the absence of reducting agent. Dissociation took place in the presence of reducing agents, which indicated that the subunits had become covalently linked by disulfide linkages. Proteins extracted from purified crosslinked 70 S ribosomes were first fractionated by polyacrylamide/urea gel electrophoresis. The proteins from sequential slices of these gels were analyzed by two-dimensional polyacrylamide/sodium dodecyl sulfate diagonal gel electrophoresis. Monomeric proteins derived from crosslinked dimers appeared below the diagonal containing non-crosslinked proteins, since the second electrophoresis, but not the first, is run under reducing conditions to cleave the crosslinked species. Final identification of the proteins in each dimer was made by radioiodination of the crosslinked proteins, followed by two-dimensional polyacrylamide/urea gel electrophoresis in the presence of non-radioactive total 70 S proteins as markers. This paper describes the identification of 23 protein dimers that contained one protein from each of the two different ribosomal subunits. The proteins implicated must have some part of their structure in proximity to the other ribosomal subunit and are therefore defined as “interface proteins”. The group of interface proteins thus defined includes 50 S proteins that are part of the 5 S RNA: protein complex and 30 S proteins at the initiation site. Correlations between the crosslinked interface proteins and other functional data are discussed.     

Waterlogging tolerance in the tribe Triticeae: the adventitious roots of Critesion marinum have a relatively high porosity and a barrier to radial oxygen loss

Nine species from the tribe Triticeae – three crop, three pasture and three ‘wild’ wetland species – were evaluated for tolerance to growth in stagnant deoxygenated nutrient solution and also for traits that enhance longitudinal O₂ movement within the roots. Critesion marinum (syn. Hordeum marinum) was the only species evaluated that had a strong barrier to radial O₂ loss (ROL) in the basal regions of its adventitious roots. Barriers to ROL have previously been documented in roots of several wetland species, although not in any close relatives of dryland crop species. Moreover, the porosity in adventitious roots of C. marinum was relatively high: 14% and 25% in plants grown in aerated and stagnant solutions, respectively. The porosity of C. marinum roots in the aerated solution was 1·8–5·4‐fold greater, and in the stagnant solution 1·2–2·8‐fold greater, than in the eight other species when grown under the same conditions. These traits presumably contributed to C. marinum having a 1·4–3 times greater adventitious root length than the other species when grown in deoxygenated stagnant nutrient solution or in waterlogged soil. The length of the adventitious roots and ROL profiles of C. marinum grown in waterlogged soil were comparable to those of the extremely waterlogging‐tolerant species Echinochloa crus‐galli L. (P. Beauv.). The superior tolerance of C. marinum, as compared to Hordeum vulgare (the closest cultivated relative), was confirmed in pots of soil waterlogged for 21 d; H. vulgare suffered severe reductions in shoot and adventitious root dry mass (81% and 67%, respectively), whereas C. marinum shoot mass was only reduced by 38% and adventitious root mass was not affected.     

Biofortification of crops with seven mineral elements often lacking in human diets – iron, zinc, copper, calcium, magnesium, selenium and iodine

The diets of over two-thirds of the world's population lack one or more essential mineral elements. This can be remedied through dietary diversification, mineral supplementation, food fortification, or increasing the concentrations and/or bioavailability of mineral elements in produce (biofortification). This article reviews aspects of soil science, plant physiology and genetics underpinning crop biofortification strategies, as well as agronomic and genetic approaches currently taken to biofortify food crops with the mineral elements most commonly lacking in human diets: iron (Fe), zinc (Zn), copper (Cu), calcium (Ca), magnesium (Mg), iodine (I) and selenium (Se). Two complementary approaches have been successfully adopted to increase the concentrations of bioavailable mineral elements in food crops. First, agronomic approaches optimizing the application of mineral fertilizers and/or improving the solubilization and mobilization of mineral elements in the soil have been implemented. Secondly, crops have been developed with: increased abilities to acquire mineral elements and accumulate them in edible tissues; increased concentrations of 'promoter' substances, such as ascorbate, β-carotene and cysteine-rich polypeptides which stimulate the absorption of essential mineral elements by the gut; and reduced concentrations of 'antinutrients', such as oxalate, polyphenolics or phytate, which interfere with their absorption. These approaches are addressing mineral malnutrition in humans globally.     

Variability of Wax Ester Fermentation in Natural and Bleached Euglena gracilis Strains in Response to Oxygen and the Elongase Inhibitor Flufenacet

ABSTRACT. Euglena gracilis is able to synthesize adenosine triphosphate under anaerobic conditions through a malonyl-independent fatty acid synthesis leading to wax ester fermentation. Mitochondrial fatty acid synthesis uses acetyl-CoA and propionyl-CoA as C2- and C3-donors for de novo synthesis of even- and odd-numbered fatty acids, respectively. Euglena' s wax ester fermentation has only been described in the E. gracilis strain 1224-5/25 Z. Here we investigate eight E. gracilis strains isolated in 1932–1958 from different localities in Europe and two bleached substrains of E. gracilis 1224-5/25, obtained by treatment with streptomycin and ofloxacin, and examine their anaerobic growth, wax ester fermentation, and wax ester composition. Under ambient oxygen levels, all strains accumulated wax esters in concentrations between 0.3% and 3.5% of the dry weight, but the strains revealed marked differences in wax ester accumulation with respect to anaerobic growth. Most fermenting strains tested showed increased wax ester synthesis under anaerobic conditions as well as the increased synthesis of odd-numbered fatty acids and alcohols suggesting an activation of the mitochondrial fatty acid biosynthesis pathway. Addition of the elongase inhibitor flufenacet to the growth medium specifically reduced the accumulation of odd-numbered fatty acids and alcohols and tended to increase the overall yield of anaerobic wax esters.     

利用FRET技术研究Hepcidin 和Fpn 相互作用

铁是生命必需的微量元素,ferroportin(Fpn)是小肠吸收细胞铁释放的重要蛋白。新近发现肝脏分泌的抗菌多肽 hepcidin 具有调节肠铁吸收的重要作用,但目前尚缺少Fpn和hepcidin发生作用的实验依据。应用荧光共振能量转移技术(fluorescence resonance energy transfer ,FRET)对hepcidin和Fpn之间的作用关系进行了深入研究。首先进行了hepcidin-CF P融合蛋白表达载体的构建及表达鉴定;然后对含YFP,Fpn-YFP基因动物细胞表达载体的构建、表达和FRET检测。实验结果证实hepcidin和Fpn之间存在直接的相互作用,并发现两种蛋白发生相互作用后hepcidin也在细胞质中有分布。为临床治疗铁代谢紊乱性疾病提供了新的治疗策略和重要理论依据。     

Conjugal transfer of aminoglycoside and macrolide resistance between Enterococcus faecium isolates in the intestine of streptomycin-treated mice

The purpose was to study conjugal transfer of resistance genes between a multi-resistant Enterococcus faecium isolate and a sensitive E. faecium isolate. Co-transfer of erm(B)-Tn5405-like element and aac(6')-Ie-aph(2')-Ia was obtained in both in vivo and in vitro. Plasmid profiles and Southern blots showed that both the erm(B)-Tn5405-like element and aac(6')-Ie-aph(2')-Ia were placed on the same large plasmid (>147 kb). These data show to our knowledge the first co-transfer of the erm(B)-Tn5405-like element and aac(6')-Ie-aph(2')-Ia. The in vivo study also indicates that transfer of resistance genes between enterococci might occur under natural conditions in the gut of animals.     

Effects of penicillin on the synthesis of membrane proteins of Chlamydia trachomatis LGV2 serotype

Abstract Actinobacillus (Hemophilus) pleuropneumoniae type strain 4074, serotype 1, secretes a potent hemolysin. This hemolysin is thermolabile and inactivated by proteinase K. We have purified the hemolysin to homogeneity and characterized it as a protein of 105 kDa by SDS-Polyacrylamide gel electrophoresis. Using a calibrated gel filtration column, the active hemolysin was identified as a monomer of the 105 kDa polypeptide. This hemolysin is an acid protein with an isoelectric point at p I 4.3.     

Metal compounds for the treatment of parasitic diseases

The cysteine proteases of the trypanosomatid parasitic protozoa have been validated as targets for chemotherapy of Chagas’ disease and leishmaniasis. Metal complexes of gold, platinum, iridium, palladium, rhodium and osmium have been reported to have activity against a variety of trypanosomatids, but the molecular target of these compounds has not been defined. The activity of gold(III) and palladium(II) cyclometallated complexes, and oxorhenium(V) complexes against mammalian and parasitic cysteine proteases was investigated. All gold(III) complexes (1-6) inhibited cathepsin B with IC₅₀ values in the range of 0.2-1.4 μM. Of the six palladium compounds, aceto[2,6-bis[(butylthio-κS)methyl]phenyl-κC]-, (SP-4-3)-palladium(II) (11) was the most potent inhibitor of cathepsin B with an IC₅₀ of 0.4 μM. A clear structure-activity relationship was observed with the oxorhenium(V) complexes with chloro[2,2′-(thio-κS)bis[ethanethiolato-κS)]] oxorhenium(V) (16) being the most potent inhibitor of cathepsin B with an IC₅₀ of 0.009 μM. Six complexes were further tested against the parasite cysteine proteases, cruzain from T. cruzi, and cpB from L. major; the most potent inhibitors were the two rhenium complexes (2(1H)-pyridinethionato-κS²)[2,6-bis[(mercapto-κS)methyl]pyridine-κN¹] oxorhenium(V) (15) and chloro[2,2′-(thio-κS)bis[ethanethiolato-κS)]] oxorhenium(V) (16). The compounds were also evaluated in assays for parasite growth. Two oxorhenium(V) compounds ((p-methoxyphenylthiolato-S)[2,6-bis[(mercapto-κS)methyl]pyridine-κN¹] oxorhenium(V) (14) and (methanethiolato)[2,2′-(thio-κS)bis[ethanethiolato-κS)]] oxorhenium (V) (18)) and the palladium compound 11 inhibited T. cruzi intracellular growth, and compound 11 inhibited promastigote growth in three Leishmania species. In conclusion this preliminary data indicates that metal complexes targeted at parasite cysteine proteases show promise for the treatment of both Chagas’ disease and leishmaniasis.     

Materials Flow Analysis of the Italian Economy

This article analyzes the mass of the materials that flowed through the Italian economy during 1994 and compares the results with a similar analysis of Germany, Japan, the Netherlands, and the United States published by a collaboration headed by the World Resources Institute. In order to perform this comparison, we have evaluated the mass of the materials produced within the country and the mass of the imported materials and commodities. For the domestic production, imports and exports, we have also evaluated the mass of the materials that accompany—as "hidden flows"—each physical flow.
Our analysis indicates that, in 1994, Italy experienced total material requirements (TMR) of 1,609 million metric tons (Mt), of which 727 Mt was used as direct material input (DMI). A comparison with other developed countries shows that the TMR and DMI flows, measured in mass per person and in mass per GDP unit, are, in Italy, lower than the corresponding figures evaluated for the United States, Germany, and the Netherlands. An interpretation of these results is presented. The analysis may give information useful for environmental considerations, although the limits of such an approach are made clear.