葡萄糖苷酶在毕赤酵母中的重组表达及一步纯化

从绿色木霉中克隆了葡萄糖苷酶bg基因,构建入表达载体pPIC9k-His6中,然后在AOX1启动子的控制下,在毕赤酵母GS115菌株中表达. 在5 L发酵罐中发酵120 h,重组P. pastoris Mut+菌株湿重达360.6 g/L,葡萄糖苷酶浓度和酶活分别为2.1 mg/mL和73.5 U/mL. 经亲和层析一步纯化后,得到了电泳纯的葡萄糖苷酶. HPLC分析显示其纯度为95.6%,比酶活为71.9 U/mg. 纯化过程酶得率为73.6%,纯化倍数为42.6. 纯酶的等电点为5.0,最适温度为50℃,最适pH为6.5. 金属离子Ag+, Ca2+, Cu2+, Fe2+及SDS对葡萄糖苷酶活性有抑制作用,而Mg2+, Mn2+, K+能增强葡萄糖苷酶活性,其中1 mol/L Mg2+能使酶活提高20%.     

argE基因的表达优化及Zn2+添加时间和浓度的影响机制分析

确定了目的基因argE在重组菌BL21(DE3)-pET22b-argE中的表达位置,研究了Zn2+对重组菌生长及表达产物活性的影响,并分析了影响机制. 结果表明,argE可在重组菌中高效表达,表达产物N-乙酰鸟氨酸脱酰基酶大多以不可溶的包涵体形式存在,只有少量为有活性的可溶性表达. 1.0 g/L的Mg2+对重组菌的生长及酶活有明显促进作用. Zn2+加入时机及加入量不同,影响结果也不同. 发酵起始加入Zn2+严重抑制菌的生长及酶活,而在1.0%乳糖诱导2.5 h后加入则可解除生长抑制并提高酶活. SDS-PAGE电泳及活力测定证实Zn2+参与形成酶的催化中心,对酶的表达量没有影响.     

一种铜锌超氧化物歧化酶突变体的原核表达、纯化及酶活性测定

目的原核表达、纯化一种铜锌超氧化物歧化酶(Cu,Zn-SOD)突变体,并检测其酶活性。方法将经热力学优化的Cu,Zn-SOD突变体基因亚克隆至pET-28a载体中,构建重组表达质粒SOD1-pET-28a,转化入大肠埃希菌BL21(DE3)中,IPTG诱导表达;在500 mL摇瓶培养条件下,通过改变加入Cu、Zn离子的量优化诱导表达条件;表达的重组蛋白经初步纯化后,测定酶活性。结果重组表达质粒SOD-pET-28a经双酶切及测序证明构建正确;表达的重组蛋白相对分子质量约为15 000,主要以可溶性形式表达;工程菌株在500 mL摇瓶培养条件下培养至10 h左右,A₆₀₀至3. 5左右时加入终浓度为0. 1 mmol/L IPTG、0. 2 mmol/L ZnCl₂、0. 5 mmol/L CuSO4,可获得最佳表达量(29. 2%);初步纯化获得的重组蛋白纯度为97. 32%,比活为5. 08×103U/mg。结论成功利用大肠埃希菌表达系统表达了一种经热力学优化的Cu,Zn-SOD突变体,初步纯化的突变体具有较好的酶活性,为进一步摸索大规模生...     

金属离子对纤维素酶内切酶和外切酶活性的影响

金属离子可以影响纤维素内、外切酶的活性,且不同金属离子对其活性影响程度也不同。据此,本实验选取三种纤维素酶进行单因素实验,对其内、外切酶活性进行研究。实验结果表明,当金属离子的质量浓度为:K+0.9 mg/mL,Ca2+0.3 mg/mL,Zn2+1.2 mg/mL,Cu2+1.2 mg/mL,能较好地提高纤维素内切酶活;而Mg2+﹑Ca2+﹑Ba2+﹑Cu2+﹑Zn2对纤维素外切酶活都有抑制作用;Al3+对纤维素内、外切酶活都有抑制作用;不同价态阴离子对纤维素内、外切酶活影响不大。     

以香豆素为母体的荧光探针的合成、离子识别及拟合计算研究

设计合成了含噻唑和腙结构的香豆素类钴离子荧光探针分子CCo,其结构用1HNMR和13CNMR进行了表征,考察了其光谱性能和电化学性能,并对其结构进行了拟合计算研究。CCo在常见金属离子(Cd2+、Co2+、Na+、Mn2+、Fe3+、Pb2+、Hg2+、Cu2+、Zn2+、Cu+、K+、Mg2+、Ag+、Ni2+、Cr3+)中能够选择性地识别Co2+。滴加Co2+后探针吸收光谱红移60 nm,荧光光谱蓝移75 nm。探针分子溶液在三电极系统及四丁基高氯酸胺作电解质下能用于Hg2+的检测。通过拟合计算进一步验证了探针吸收光谱峰值与实验值一致。     

腾冲菌脂肪酶的表达纯化及酶学性质的表征

将来自腾冲菌的脂肪酶(Lip A)基因(lip A)克隆到大肠杆菌表达载体p ET28a(含T7启动子)和p Trc99A(含Trc启动子)中,转入大肠杆菌表达,发现Trc启动子更适合Lip A的表达。通过热处理和DEAE-Sepharose阴离子柱纯化过程,重组Lip A得到纯化,比酶活达到1.9U/mg,重组Lip A分子量为42k Da。重组Lip A在80℃、p H 4.5时酶活最高,经85℃保温2h,酶活保持60%以上;在p H值4.0~6.0之间,Lip A具有较好的稳定性;Cu2+和Zn2+对酶活力分别有38.9%和69.2%的抑制作用,Mn2+、Co2+和Tween-20对该酶有较大的激活作用;以p-nitrophenyl-laurate(p-NP-C12)为底物时,该酶的Km值为1.5mmol/L,kcat为34.5s-1。     

酶法转化制备L-瓜氨酸

利用粪链球菌精氨酸脱亚胺酶转化L-精氨酸制备L-瓜氨酸。考察了菌龄、转化温度等多种因素对精氨酸脱亚胺酶活力的影响。酶法制备L-瓜氨酸的最适工艺条件是:菌体发酵时间20 h,转化温度37℃,转化液起始pH为6.0,ρ〔十六烷基三甲基溴化铵(CTAB)〕=0.3 g/L使酶活提高了414%。c(Co2+)=10-3mol/L使酶活增高约50%,c(Cu2+)=10-2mol/L或c(Zn2+)=10-2mol/L使酶活下降约50%。Ca2+、Mg2+和Mn2+对酶活影响较小。     

受主掺杂对钛酸锶氧敏传感器的影响

简单介绍了汽车用氧传感器,总结Mg2+,Fe3+,Al3+,Li+,Ag+,Mn2+,Cu2+,Ni2+等受主离子掺杂对钛酸锶氧敏性能的影响.综合比较铁、镁掺杂钛酸锶效果较好.     

香豆素类荧光增强型Cu2+荧光分子探针

设计合成了含酰肼和亚胺结构的双香豆素类Cu2+荧光分子探针CCu,其结构用1HNMR、IR、HRMS和元素分析进行了表征,并考察了其光谱性能。CCu在常见碱金属离子(K+,Na+)、碱土金属离子(Ca2+,Mg2+)、过渡金属及重金属离子(Cd2+,Ag+,Fe3+,Pb2+,Hg2+,Cr3+,Co2+,Ni2+,Cu2+,Zn2+)中能够专一性地识别Cu2+。滴加Cu2+后引起吸收光谱蓝移38 nm,荧光光谱蓝移17 nm,荧光增强8.2倍,溶液荧光由无色变为蓝色。CCu对pH不敏感具有较低的pKa值(3.32±0.07),可在近中性和弱碱性范围内对Cu2+进行检测。     

过氧化氢氧化玉米秸秆酶解残渣制备螯合微肥的研究

利用过氧化氢在不同条件下对玉米秸秆制备生物乙醇的酶水解残渣进行氧化改性,氧化产物分别与Fe2+、Cu2+、Zn2+、Mn2+四种离子进行螯合反应制备微肥。探讨了不同氧化条件对金属离子螯合率的影响,并确定了适宜的氧化工艺条件:过氧化氢用量20%,温度70℃,pH值为11,时间60 min,此时残渣溶出率为32.5%,氧化产物对Fe2+、Cu2+、Zn2+、Mn2+四种金属离子的总螯合率分别为5.0%,4.5%,3.9%,4.0%。通过正交试验分析,氧化条件对酶解残渣的溶出率和总螯合率的影响主次顺序为:pH值,过氧化氢用量,温度。     

Precipitate Foam Flotation of Copper,Nickel, Cobalt,and Manganese with LIX Reagents

Abstract

The precipitate flotation of copper, nickel, cobalt, manganese, zinc, and cadmium with LIX65 and LIX63 (hydroxyoxime compounds) and carrier surfactants hexadecyltrimethylammonium bromide and TX100 was investigated. The effects of pH and possibly interfering ions (sulfate, phosphate, EDTA) were studied. LIX65 was found to give good results with copper, nickel, cobalt, and manganese; LIX63 was less satisfactory.     

Treatment of wastewater containing copper,zinc, nickel and cobalt using Duolite ES‐467

Duolite ES‐467 was used to treat wastewater containing heavy metal ions. Sorption experiments were carried out at varying pH values, agitation speeds, reaction times, and metal ion and sorbent concentrations. Each of the parameters affects the sorption behaviour of individual metal ions. Copper sorption was greater compared with other metal ions such as zinc, nickel and cobalt. The presence of other metal ions affects copper sorption. Equilibrium isotherm curves were developed. These were used to predict that the metal ion concentration would be reduced from 100 to less than 1 mg dm^?3. Fixed bed tests were conducted to investigate the efficiency of Duolite ES‐467 for the selective removal of copper ions from multi‐metal solutions. Breakthrough curves were obtained using Duolite ES‐467 for solutions containing copper, nickel and copper, zinc, nickel and cobalt. Elution studies were also carried out using sulfuric acid. © 2002 Society of Chemical Industry     

铁氧体法去除废水中的镍、铬、锌、铜离子

采用铁氧体法处理含镍、铬、锌、铜的废水,研究了pH及硫酸亚铁投加量对重金属离子去除效果的影响.对于镍、锌、铜离子,最佳絮凝pH分别为8.00～9.80、8.00～10.50和10.00,投加的亚铁离子与其摩尔比均为2～8;六价铬的最佳还原pH为4.00～5.50,最佳絮凝pH则为8.00～10.50,最佳投料比为20....     

Effect of metallic impurities on the gasification of graphite in water vapor and hydrogen

The catalytic effects of metallic impurities on the reactivity of graphite towards water vapor and hydrogen in the temperature range 25° to 1100°C have been investigated as a function of the oxidation state of the impurity. Iron, cobalt and nickel are active catalysts for the former reaction between 600° and 1000°C when the metal is kept in the reduced state by means of added hydrogen. Motion of the metallic particles on the basal plane surface of the graphite during the reaction leads to the formation of channels which with the smallest catalyst particles are oriented mostly in the 〈112̄0〉 directions. Vanadium and molybdenum are weak catalysts under these conditions, whereas copper, zinc, cadmium, silver, chromium, manganese and lead are inactive. When hydrogen is absent so that the metal remains in the oxidized state, the catalytic activity of all these impurities is low or negligible. The reaction of graphite with dry hydrogen occurs less readily but is again strongly catalyzed by metallic iron, cobalt and nickel. Manganese, chromium, molybdenum and vanadium show a slight catalytic effect in the hydrogenation reaction at temperatures around 1000°C, whereas copper, zinc, cadmium, silver and lead are inactive.     

Purification of nickel sulphate solutions containing iron,copper, cobalt,zinc and manganese

Acidic nickel-bearing solution containing iron, cobalt, manganese, zinc and copper was processed through a solvent extraction and precipitation technique to obtain a pure nickel sulphate solution. Iron was extracted using 0.2M Cyanex-272 (partially neutralised) as the extractant. Stripping of iron from the loaded organic has also been studied. After iron recovery through solvent extraction the raffinate still contained 0·25 g dm^?3 of iron which was quantitatively separated by a lime precipitation technique. During this iron precipitation there was no loss of cobalt and nickel but copper, manganese and zinc were coprecipitated to some extent. From the iron-free nickel sulphate solution the other impurities were extracted using the same extractant (Cyanex-272) in a single stage. The metal ions from the loaded organic were stripped using a 0·5% (v/v) H₂SO₄ solution in a single stage. The entire operation needs only seven stages: two stages for iron extraction, three stages for iron stripping from the loaded organic, and one stage each for extraction and stripping of other impurities. In the entire operation the loss of nickel was less than 0·5%.     

Sorption isotherms of metal ions onto an N-(2-sulfoethyl)chitosan-based material from single- and multi-component solutions

The scope of work is to study the mutual influence of metal ions during their sorption by sulfoethylated chitosan. The sorption isotherms of metal ions from single- and multi-component solutions are obtained. The sorption capacity of the sorbent towards Ag(I) and Cu(II) is revealed to be 1.63 and 1.41 mmol/g in single-, and 1.40 and 0.85 mmol/g in five-component solution. By comparing the affinity parameter and capacity of sulfoethylated chitosan towards ions in single- and multi-component solutions, it is concluded that Ag(I) and Cu(II) ions suppress the sorption of cobalt(II), nickel(II), zinc(II), cadmium(II), magnesium(II), calcium(II), strontium(II), barium(II), manganese(II) and lead(II).     

Reduction electrochemique des spinelles Zn NixMn2-xO4 (1 χ 0)

It is demonstrated through the electrochemical reduction of zinc manganese spinels that it is possible to increase the cathodic reduction by replacement of manganese ions by nickel cations. The best performance is showed by ZnNi_0.5Mn_1.5O₄ spinel. The reduction reaction occur on octahedral sites formed by Mn⁴⁺ ions associated with Mn³⁺ ions. The electrochemical reaction is likely to occur by the redox on solid state between nickel and manganese ions.     

Comparison of Biosorption of Nickel (II) and Copper (II) Ions from Aqueous Solution by Sphaeroplea Algae and Acid Treated Sphaeroplea Algae

Abstract

Biosorption of nickel (II) and copper (II) ions from aqueous solution by dead sphaeroplea algae in natural and acid treated forms were studied as a function of concentration, pH, and adsorbent dose. The optimum pH for nickel (II) and copper (II) biosorption was found to be 6.0 and 4.0 respectively. The metal ion uptake increased with initial metal ion concentration studied up to 500 mg/L. Both the Freundlich and Langmuir adsorption models could fit the equilibrium data. The adsorption reasonably fitted the Lagergren kinetic model. Further the biomass was characterized by FTIR spectra. Surface area values are measured to be 0.9 and 2.1 m²/g for natural and acid treated forms respectively. The maximum adsorption capacity was found to be 3.40, 4.15 mmol/g for nickel (II) and 2.21, 3.41 mmol/g for copper (II) in natural and acid treated forms respectively.     

氨配合法制活性氧化锌除杂净化过程研究

本文对氨水－碳酸氨作浸取液以粗氧化锌为原料制活性氧化锌的除杂净化过程进行了研究。氨配合法生产活性氧化锌深度除杂净化中，以H2O2作氧化剂深度除锰，以（NH4）2S为沉淀除杂剂深度除铜，使低品位的原料浸取液得以净化。     

Enzymatic activity of Campylobacter jejuni hippurate hydrolase

The hippurate hydrolase enzyme of Campylobacter jejuni was expressed in Escherichia coli as a six-histidine-tagged fusion protein. The purified recombinant enzyme was characterized to gain an understanding of the structure and activity of the hippurate hydrolase. The recombinant enzyme had a native molecular mass of 193+/- 11 kDa a reduced molecular mass of 42.4+/- 0.8 kDa, and possessed 1.98+/- 0.68 molecules of zinc per enzyme subunit molecule, suggesting that it was a homotetramer with two associated zinc ions. The enzyme was a metallocarboxypeptidase that was sensitive to silver, copper and ferrous ions, and displayed optimal activity at pH 7.5 and 50 degrees C. It hydrolyzed carboxypeptidase substrates in vitro, displaying its highest activity against N-benzoyl-linked small aliphatic amino acids. A high proportion of the enzyme structure consisted of highly ordered alpha-helix and beta-sheet sequences. An alignment of the amino acid sequence of the hippurate hydrolase enzyme with those of related enzymes with similar activities revealed several conserved amino acids, which might be involved in enzyme catalysis or metal ion binding for the enzyme. Site-directed mutagenesis of the recombinant enzyme demonstrated that the Asp(76), Aps(104), Glu(134), Glu(135), His(161) and His(356) positions were important for the catalytic activity of the enzyme.