Purification, characterization and primary structure of a chymotrypsin inhibitor from Naja atra venom

A chymotrypsin inhibitor, designated NA-CI, was isolated from the venom of the Chinese cobra Naja atra by three-step chromatography. It inhibited bovine (x-chymotrypsin with a K-i of 25 nM. The molecular mass of NA-CI was determined to be 6403.8 Da by matrix-assisted laser-desorption ionization time-of-flight (MALDI-TOF) analysis. The complete amino acid sequence was determined after digestion of S-carboxymethylated inhibitor with Staphylococcus aureus V8 protease and porcine trypsin. NA-CI was a single polypeptide chain composed of 57 amino acid residues. The main contact site with the protease (PI) has a Phe, showing the specificity of the inhibitor. NA-CI shared great similarity with the chymotrypsin inhibitor from Naja naja venom (identities = 89.5%) and other snake venom protease inhibitors. (C) 2003 Elsevier Inc. All rights reserved.

Separation and purification of ovulation-inducing factors in the seminal plasma of the Bactrian camel (Camelus bactrianus)

Ovulation in the Bactrian camel depends upon ovulation-inducing factors in the seminal plasma. The present study was conducted to isolate and purify the bioactive fractions from the seminal plasma of these camels. The seminal plasma was fractionated by anion-exchange chromatography, and six fractions were obtained. The bioactive potential of each fraction was estimated from its effect on rat pituitary tissue cultured in vitro and by the effect of an intramuscular injection of the fraction into female camels in vivo. Both the third fraction (F3) and the fifth fraction (F5) stimulated the release of LH in vitro and in vivo. In addition, female camels ovulated within 48 h after intramuscular injection of F3. However, neither F3 nor F5 had any significant effect on the secretion of FSH, either in vitro or in vivo. When F3 was further fractionated into four subfractions, the third subfraction (F3-3) still stimulated the in vitro release of LH, but not of FSH. An attempt to further purify the ovulation-inducing factors in F3-3 failed owing to the similarity of the molecular characters.

Purification, characterization and biological activities of the L-amino acid oxidase from Bungarus fasciatus snake venom

L-Amino acid oxidases (LAAOs) are widely distributed in snake venoms, which contribute to the toxicity of venoms. However, LAAO from Bungarus fasciatus (B. fasciatus) snake venom has not been isolated previously. In the present study, LAAO from B. fasciat

Expression, purification and characterization of recombinant Jerdonitin, a P-II class snake venom metalloproteinase comprising metalloproteinase and disintegrin domains

Jerdonitin is a P-II class snake venom metalloproteinase comprising metalloproteinase and disintegrin domains. In this study, we established a high-level expression system in Pichia pastoris and developed a purification strategy for the recombinant Jerdonitin. This recombinant Jerdonitin degraded fibrinogen at a level of activity comparable with its wild type. The effects of recombinant Jerdonitin on inhibiting ADP-induced human platelet aggregation were in a dose-dependent manner with an IC50 of 248 nM. In addition, we reported here that Jerdonitin can significantly inhibit the growth of several cell lines, including human liver cancer cells (Bel7402), human leukemia cells (K562) and human gastric carcinoma cells (BGC823). This study offers recombinant Jerdonitin that will be valuable for further functional and structural studies of Jerdonitin. (C) 2009 Elsevier Ltd. All rights reserved.

Estimation of the natural purification rate of a eutrophic lake after pollutant removal

Pollution resulting from increased human activities is threatening Lake Donghu, its effects being characterized by serious eutrophication. A steady increase of phosphorus loading is the most important factor of the lake eutrophication. Pollution external control projects are being implemented and will be accomplished before the year 2010. In order to predict the restoration rate by the lake's self-purification after the projects of external control, a model of predicting the removal rate of total phosphorus (TP) from lake water is developed, and a brief method of estimating the release and export rate of sediment phosphorus is suggested. Results show that, on the premise of external loading fully controlled. The restoration needs about 55 years or more. Obviously, the great P pool in the sediment will be a limiting factor of preventing the improvement of water quality after the external loading is under control. Based on the estimation we conclude that after the external control projects before 2010, in order to restore the lake in a few years, although highly cost, the first step must be the sediment dredging to remove internal loading. The second step is diverting water of River Changjiang into the lake to accelerate the improvement of lake water. Otherwise, removal of pollutant sources will become meaningless. (c) 2006 Elsevier B.V. All rights reserved.

Purification and characterization of hydrosoluble components from the sap of Chinese lacquer tree Rhus vernicifera

Continuous gradient elution chromatography (CGEC) was employed to purify and separate enzymes and polysaccharides from the sap of Rhus vernicifera Chinese lacquer tree. There are three different molecules with laccase enzyme activity. Two are enzymes of each other (L1, and L2), whereas the third (RL) is an entirely separate entity. Two polysaccharides (GP1 and GP2) were also found. The Rhus laccase (RL), and isoenzymes L1 and L2, have peak molecular masses of 109,100, 120,000, 103,000 respectively; each has four copper atoms per molecule, and the pI values were 8.2, 8.6, and 9.1, respectively. The structure of the laccases was studied by Fourier-transform infrared (FT-IR) and Matrix-assisted laser desorption/ionization time-of flight (MALDI-TOF) mass spectrometry. The typical amide I (1646 cm(-1)) and amide II (1545 cm(-1)) bands were observed. The results from MALDI-TOF were similar to those from CGEC, but the molecular mass from the MALDI-TOF was significantly different from that obtained from sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). (c) 2006 Elsevier B.V. All rights reserved.

Root zone microbial populations, urease activities, and purification efficiency for a constructed wetland

In order to investigate the effects of microorganisms and their urease activities in macrophytic root zones on pollutant removal, four small-scale plots (SSPs) of vertical/reverse-vertical flow wetlands were set up to determine: a) the relationship between the abundance of microorganisms in the root zones and water purification efficiency; and b) the relationship between urease activities in the root zones and pollutant removal in a constructed wetland system. Total numbers of the microbial population (bacteria, fungi, and actinomyces) along with urease activities in the macrophytic root zones were determined. In addition, the relationships between microbial populations and urease activities as well as the wastewater purification efficiencies of total phosphorus (TP), total Kjeldahl nitrogen (TKN), biochemical oxygen demand in 5 days (BOD5), and chemical oxygen demand (COD) were also analyzed. The results showed that there was a highly significant positive correlation (r = 0.9772, P < 0.01) between the number of bacteria in the root zones and BOD5 removal efficiency and a significant negative correlation (r = -0.9092, P < 0.05) between the number of fungi and the removal efficiency of TKN. Meanwhile, there was a significant positive correlation (r = 0.8830, P < 0.05) between urease activities in the root zones and the removal efficiency of TKN. Thus, during wastewater treatment in a constructed wetland system, microorganism and urease activities in the root zones were very important factors.

Roles of substrate microorganisms and urease activities in wastewater purification in a constructed wetland system

A vertical/reverse-vertical flow constructed wetland system was set up in Wuhan, China, to study its treatment efficiency of polluted lake water. The numbers of substrate microorganisms and urease activities in the substrate of the constructed wetland were determined by plate counts and colorimetric analysis, respectively. The removal efficiencies of biochemical oxygen demands (BOD5). chemical oxygen demands (COD), total phosphorus (TP), total Kjeldahl nitrogen (TKN), and total suspended solids (TSS) were measured by EPA approved methodology. The results showed there were significant positive correlations (P < 0.05) between numbers of microorganism in the substrate and removal rates of TKN and CODCr. Meanwhile, there was significant positive correlation (P < 0.05) between urease activities and removal efficiencies of TKN and negative correlation between urease activities and removal efficiencies of BOD5. Substrate microorganisms and urease activities played key factors during purification processes and they could be utilized as indicator of wastewater treatment performances in the constructed wetland system. (C) 2004 Elsevier B.V. All rights reserved.

Purification and characterization of White Spot Syndrome Virus (WSSV) produced in an alternate host: crayfish, Cambarus clarkii

Penaeid shrimp is the natural host of White Spot Syndrome Virus (WSSV) that can cause high mortality in the infected hosts. Attempts to obtain sufficient amounts of purified intact WSSV for characterization have been unsuccessful. Using crayfish, Cambarus clarkii as a proliferation system, a large amount of infectious WSSV was reproduced and intact WSSV viral particles were purified with a new isolation medium by ultra-centrifugation. Purified WSSV particles were very sensitive to organic solvents and the detergent, Triton X-100. The size of the rod-shape, somewhat elliptical, intact WSSV was 110-130 x 260-350 mm with a long, tail-like envelope extension. The naked viral nucleocapsid was about 80 x 350 nm, and it possessed 15 spiral and cylindrical helices composed of 14 globular capsomers along its long axis, and a 'ring' structure at one terminus. Distinct WSSV genome DNA patterns were obtained when the purified genomic dsDNA of WSSV was digested with five different restriction enzymes (HindIII, XhoI, B(BamHI, SalI, and SacI). In addition, at least 13 major and distinct protein bands could be observed when purified intact WSSV viruses were separated by SDS-PAGE followed by Coomassie Brilliant R-250 staining. The estimated molecular weights of these proteins were 190, 84, 75, 69, 68, 58, 52, 44, 28, 27.5, 23, 19, and 16 kD, respectively. Both the 44 and 190 kD proteins were easily removed if the hemolymph from the: WSSV infected crayfish was transiently treated with 1%, Triton X-100 before it was subjected to gradient centrifugation, indicating that both of them are located on the surface of the viral envelope. These characteristics are consistent with WSSV isolated from the penaeid shrimp. (C) 2001 Elsevier Science B.V. All rights reserved.

PURIFICATION AND CHARACTERIZATION OF A KINASE SPECIFIC FOR THE SERINE-RICH AND ARGININE-RICH PRE-MESSENGER-RNA SPLICING FACTORS

Members of the SR family of pre-mRNA splicing factors are phosphoproteins that share a phosphoepitope specifically recognized by monoclonal antibody (mAb) 104. Recent studies have indicated that phosphorylation may regulate the activity and the intracellular localization of these splicing factors. Here, we report the purification and kinetic properties of SR protein kinase 1 (SRPK1), a kinase specific for SR family members. We demonstrate that the kinase specifically recognizes the SR domain, which contains serine/arginine repeats. Previous studies have shown that dephosphorylated SR proteins did not react with mAb 104 and migrated faster in SDS gels than SR proteins from mammalian cells. We show that SRPK1 restores both mobility and mAB 104 reactivity to a SR protein SF2/ASF (splicing factor 2/alternative splicing factor) produced in bacteria, suggesting that SRPK1 is responsible for the generation of the mAb 104-specific phosphoepitope in vivo. Finally, we have correlated the effects of mutagenesis in the SR domain of SF2/ASF on splicing with those on phosphorylation of the protein by SRPK1, suggesting that phosphorylation of SR proteins is required for splicing.

STUDIES ON THE PURIFICATION AND RECLAMATION OF WASTE-WATER FROM A MEDIUM-SIZED CITY BY AN INTEGRATED BIOLOGICAL POND SYSTEM

The feasibility of an inexpensive wastewater treatment system is evaluated in this study. An integrated biological pond system was operated for more than 3 years to purify the wastewater from a medium-sized city, Central China. The experiment was conducted in 3 phases with different treatment combinations for testing their purification efficiencies. The pond system was divided into 3 functional regions: influent purification, effluent upgrading and multi-utilization. These regions were further divided into several zones and subzones. Various kinds of aquatic organisms, including macrophytes, algae, microorganisms and zooplankton, were effectively cooperating in the wastewater treatment in this system. The system attained high reductions of BOD5, COD, TSS, TN, TP and other pollutants. The purification efficiencies of this system were higher than those of most traditional oxidation ponds or ordinary macrophyte ponds. The mutagenic effect and numbers of bacteria and viruses declined significantly during the process of purification. After the wastewater flowed through the upgrading zone, the concentrations of pollutants and algae evidently decreased. Plant harvesting did not yield dramatic effects on reductions of the main pollutants, though it did significantly affect the biomass productivity of the macrophytes. The effluent from this system could be utilized in irrigation and aquaculture. Some aquatic products were harvested from this system and some biomass was utilized for food, fertilizer, fodder and some other uses. The wastewater was reclaimed for various purposes.

Purification of single-walled carbon nanotubes synthesized by the catalytic decomposition of hydrocarbons

A procedure for purifying single-walled carbon nanotubes (SWNTs) synthesized by the catalytic decomposition of hydrocarbons has been developed. Based on the results from SEM observations, EDS analysis and Raman measurements, it was found that amorphous carbon, catalyst particles, vapor-grown carbon nanofibers and multi-walled carbon nanotubes were removed from the ropes of SWNTs without damaging the SWNT bundles, and a 40% yield of the SWNTs with a purity of about 95% was achieved after purification. (C) 2000 Elsevier Science Ltd. All rights reserved.

The new exploration for proton-implanted silicon: the conversion of a surface-region-purification-induced p-n junction into a p-i-n electrical structure approaching silicon on insulator

A surface-region-purification-induced p-n junction, a puzzle discovered at Brookhaven National Laboratory, in a silicon-on-defect-layer (SODL) material has been explored by carrying out various annealing conditions and subsequent measurements on electrical properties. The origin of the pn junction has been experimentally investigated. Furthermore, the p-n junction has been transformed into a p-i-n electrical structure by adding a high temperature annealing process to the previously used SODL procedure, making the SODL material approach silicon on insulator (SOI). The control of the initial oxygen amount in the silicon material is suggested to be critical for the experimental results.

基于PAR的高可信装备保障算法推演

算法被誉为计算的“灵魂”（spirit of computing），算法程序的可靠性和正确性对软件系统的可信度起着至关重要的作用。动态规划、贪心、分支限界等传统的算法设计策略缺乏有效的选择标准，一些算法形式化开发方法和工具的应用水平和范围也很有限，因此算法的可靠性已成为高可信软件系统的一个主要瓶颈。 作为军事运筹学的一个重要分支，装备保障计算主要研究军事活动中装备领域的决策优化问题，保障计算软件特别是算法的可靠性在很大程度上决定了装备保障的效率乃至军事行动的成败。利用形式化软件工程的研究成果、特别是引入算法程序的形式化开发方法，对我军通用装备保障领域中的大量算法类问题进行有效求解，有利于优化保障结构、统筹保障资源、整合保障力量、提高保障时效，促进装备整体战斗力水平的提升。 本文以高可信软件开发方法PAR为基础，面向装备保障计算的领域需求，提出了一类离散最优化问题（discrete optimization problem, DOP）的结构模型和算法推演技术，并成功推演了一系列典型的装备保障算法。本论文的主要创新性贡献如下： （1）定义了DOP的结构模型，提出了基于单点结构（Singleton）的问题分划递推策略，进而通过PAR算法推演生成问题的高效求解算法，涵盖了多种传统算法设计策略，显著提高了算法程序设计的机械化水平。 （2）提出了描述DOP分划递推过程的问题简约图（problem reduction graph, PRG）模型，并针对典型DOP结构推演得到了一组PRG构造算法模式，其中每个算法模式都涵盖了满足特定代数结构性质的一大类具体问题。 （3）阐述了PAR算法推演的范畴模型，为算法设计和重用提供了抽象而有效的方法指导。 （4）对我军通用装备保障领域具有代表性的48个问题进行了形式化的算法推演，构建了领域算法库。 （5）在PAR平台的基础上设计了装备保障算法开发平台的原型COPALM，支持DOP算法推演和重用，进而提高了相关应用软件的开发效率。