四种金属离子亲和层析纯化重组戊型肝炎病毒包涵体的效果比较

目的 选择固相金属亲和层析柱的金属离子,以优化分离纯化重组戊型肝炎病毒包涵体的条件。方法 在同一试验条件下用4种金属离子柱分离纯化目的蛋白。结果 包涵体经不同离子柱层析时目的蛋白的收获率、纯度、洗脱条件各异。结论 镍离子柱纯化效果最佳。     

pH值和盐浓度对金属螯合亲和层析分离人胰高血糖素样肽-1融合蛋白的影响

目的探讨pH值和盐浓度对金属螯合亲和层析(Immobilized metal ion affinity chromatography,IMAC)分离含His-Tag标签的人胰高血糖素样肽-1(Glucagon-like peptide-1,GLP-1)融合蛋白的影响,并确定最佳洗脱条件。方法在0.50mol/L盐浓度条件下,分别取pH6.0、7.0、7.8和9.04种缓冲液进行咪唑梯度洗脱;在合适的pH值条件下,分别取氯化钠浓度0.25、0.50和0.75mol/L3种缓冲液进行咪唑梯度洗脱。分析在不同pH值及盐浓度条件下洗脱融合蛋白所需要的咪唑浓度及回收率。结果在所考察的pH值范围内,洗脱融合蛋白所需的咪唑浓度随洗脱液pH值的升高而逐渐降低,当洗脱液pH值为7.8时,融合蛋白的分离效果最佳,大部分杂蛋白被0～40mmol/L咪唑洗脱液去除且不含融合蛋白,而绝大部分融合蛋白在咪唑浓度达80～200mmol/L时被洗出,总回收率达(83.7±1.0)%,且比例较高;在此pH值条件下,氯化钠浓度达0.75mol/L,才会影响融合蛋白的回收率,氯化钠浓度为0.25或0.50mol/L的洗脱液洗出融合蛋白的效果相近;在pH7.8,含0.25mol/L氯化钠的洗脱体系下,用50mmol/L咪唑溶液洗脱杂蛋白,200mmol/L咪唑溶液洗脱融合蛋白,融合蛋白的回收率和比例分别可达(85.2±2.0)%和(80.5±1.0)%。结论在所考察的pH值和盐浓度范围内,提高洗脱液pH值和盐浓度均可降低洗脱融合蛋白需要的咪唑量,但也会降低融合蛋白的回收率。     

壳聚糖涂层亲和层析在超氧化物歧化酶纯化中的应用

1前言亲和层析是蛋白质纯化中分离效率最高的技术之一。亲和配基有多种形式如单机、底物、活性染料、金属离子等。固定化金属离子亲和层析（Immobilizedmetalionaffinitychromatog’“PhyIMAC）是Porath在1975年首先研究的一种高效分离技术［‘3。IMAC和普通的亲和层析相比，具有一些优点，如亲和配基Cu’”、Zn’“价廉；可在高盐浓度下操作；稳定，容易再生。Sulkowke曾用IMAC技术分离纯化干扰素，一步可纯化产品几十倍【“，收率达90％。目前IMAC介质制备是先在软基质SePharose或SePhadex上用化学交联法弓卜螫合剂IDA（Imino…     

丙型肝炎病毒非结构区NS5基因片段的克隆和表达

从抗 HCV和HCVRNA均为阳性的病人血清中 ,采用RT PCR技术扩增得到HCVNS5基因片段。用含此基因片段的重组质粒pET 2 8a转化大肠杆菌BL2 1(DE3)后可高效表达NS5蛋白。该蛋白可用固定化金属离子螯合层析法 (IMAC)纯化。经Westernblot和ELISA检测结果表明 ,纯化的NS5蛋白可与丙型肝炎患者血清发生特异反应 ,在HCV感染的诊断中可能具有良好的应用前景。     

Preparation and characterization of a macroporous agarose monolith as a stationary phase in IMAC chromatography

A macroporous monolith used as stationary phase for the separation of biomolecules by immobilized metal ion affinity chromatography (IMAC), based on D5 agarose (D5) chemically modified was proposed. The characterization of physical properties was studied. Pressure drop was <0.4?MPa, being a very low value compared to other similar chromatographic supports. The adsorption/desorption process was carried out using bovine serum albumin (BSA) at pH 7.4 as a target protein. The monolith was re-used for 20 adsorption/desorption cycles and it was possible to verify that the average percentage of adsorption in all cycles was 89.65%. It was also possible to apply a model in order to obtain the kinetic adsorption constant (k_a), desorption constant (k_d) and equilibrium constant (K_e) by the proposed system. These results indicate that this system is governed by the adsorption process.     

Inhibition of angiotensin I-converting enzyme by wheat gliadin hydrolysates

A tryptic gliadin hydrolysate was fractionated into peptide fractions, which were assigned to either the central domain (CD) or terminal domains (TD) of gliadins. The domains were expected to contain amino acid (AA) sequences which, when released from the parent protein, inhibit the angiotensin I-converting enzyme (ACE), which plays a key role in regulating blood pressure. A proline (Pro) poor TD related fraction, containing the smallest peptides, showed the highest ACE inhibitory activity (IC₅₀ = 0.33 mg/ml). Additional peptidases were selected based on their in silico predicted ability to release ACE inhibitory peptides. Further hydrolysis of the tryptic hydrolysate fractions with thermolysin, Clarex, Alcalase and Esperase increased ACE inhibitory activities. Immobilised Ni²⁺-ion affinity chromatography (IMAC) purification of a TD related peptide fraction obtained by sequential hydrolysis with trypsin and thermolysin yielded a fraction with an IC₅₀ value of 0.02 mg/ml. This IMAC fraction was enriched in histidine and hydrophobic AA (Pro, Val, Ile, Leu and Phe).     

增量型模型算法控制在PMSM速度控制中的应用

针对普通PID控制系统性能受参数变化及各种不确定性影响严重等缺点,在基本模型算法控制（MAC）基础上,综合了MAC在线实时预测、优化、反馈校正的优点,在控制器中引入了积分因子组成增量型模型算法控制（IMAC）,并将该算法应用于永磁同步电机（PMSM）速度控制中。仿真结果表明,由该算法构成的控制器较普通的PID控制,在系统模型参数与实际参数严重失配的情况下,能够有效抑制各种不确定性的不良影响,且系统鲁棒性强,又可获得令人满意的动态性能。     

铜螯合亲和层析分离抗氧化活性蚕豆蛋白酶解物

采用铜离子螯合亲和层析柱对不同铜螯合能力的蚕豆蛋白酶解物组分进行分离,并探讨其抗氧化活性机制与铜螯合能力之间的关系。结果表明,金属亲和层析的最优条件为:平衡缓冲液为p H 5.0,浓度为0.05 mol/L的Na Ac-HAc;上样量为20 mg/m L的蚕豆蛋白酶解物1 m L;洗脱剂为0.04 mol/L咪唑。洗脱得到不能螯合铜的蚕豆蛋白酶解物组分F_1及能螯合铜的组分F_2,测定其总还原力、抑制羟自由基能力与铜螯合量,发现抗氧化活性的关系为F_2蚕豆蛋白酶解物F_1(P0.05),铜螯合量的关系也为F_2蚕豆蛋白酶解物(未经分离)F_1(P0.05),表明蚕豆蛋白酶解物的铜螯合活性越高,其抗氧化活性越高。     

Research on Positioning Error Compensation for Micro Milling Machine Tool

Micro milling has many advantages in fabricating three-dimensional （3D） structure in micrometer scale. The micro milling machine tool with high positioning accuracy is of great importance for getting micro structure with high profile precision and good surface quality. Meanwhile, the method of position error compensation is a good way to improve the accuracy of the micro milling machine tools. In this paper, a software method is adopted to compensate the positioning error and improve the positioning accuracy. According to error cancellation theory, the compensation values are generated and compensation tables are built to adjust the positioning error in the NC system based on Industrial Motion and Automation Control （IMAC）. The positioning accuracy of linear motor is ±0. 3 μm without backlash after compensation. In order to verify the effectiveness of compensation on the machining performance, concave spherical surfaces are processed on the micro milling machine tool. The experimental results show that the profile radius error of the spherical surface machined with compensation decreases more than 60%.     

Adsorptive removal of sulfamethoxazole with shell-core chitosan immobilized metal ion

Immobilized metal ion affinity chromatography (IMAC) for adsorptive removal of sulfamethoxazole (SMX) was investigated. Shell-core chitosan was applied as a support for metal ions and could successfully immobilize copper via ion-exchange ability of chitosan. The copper-immobilized shell-core chitosan has adsorption ability for SMX at pH = 4–6, due to the complexation between copper on the adsorbent and sulfonamide group of the SMX. The adsorbent was also applicable for chromatographic operation for removal of SMX. The SMX adsorbed was successfully eluted with deionized water, although a small amount of copper was also eluted from the adsorbent together with the SMX.