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41.
42.
高效弱阳离子交换色谱法对脲还原变性溶菌酶的折叠研究 总被引:2,自引:0,他引:2
用高效弱阳离子交换色谱(HPWCX)对脲还原变性溶菌酶(Lys)进行了复性研究. 在流动相中脲浓度固定为4.0 mol•L-1和选用对天然态蛋白有稳定作用的硫酸铵为盐或置换剂时, 在蛋白浓度为15.0~50.0 mg•mL-1时, HPWCX法比稀释法活性回收率高. 为了提高Lys的质量及活性回收率对所用色谱条件进行了优化研究, 当蛋白起始浓度为20.0 mg•mL-1时, Lys的质量回收率和活性收率分别为97.8%和95.4%. 表明此种方法简便且有可能对其他还原变性蛋白的复性具有通用性. 相似文献
43.
Z及logI对离子交换色谱中脲变溶菌酶分子构象变化的表征 总被引:3,自引:0,他引:3
以溶菌酶(Lys)为目标蛋白,用计量置换保留理论(SDT-R)中的参数Z和logI对弱阳离子交换色谱(WCX)中“准天然态”和脲变还原与非还原的两种变性状态的Lys的分子构象变化进行了表征。发现在流动相中含有脲时,蛋白的保留仍服从SDT理论,可以准确测定在特定脲浓度条件下Lys的Z及logI值。结果表明,3种分子构象状态的Lys的Z值均随脲浓度改变呈现不连续变化;“准天然态”Lys在同一脲浓度条件下的Z值比变性状态的大,logI比变性状态的小,而非还原变性态和“准天然态”Lys的Z和logI值比较接近。还 相似文献
44.
ABSTRACT We study by computer simulation the dynamics of hydration water in solution with lysozyme upon approaching the glassy state of water. We calculate the self-density correlation function at different wavelengths to test the Mode Coupling Theory (MCT) of glassy dynamics at different length scales. The results show a strict and clear relation of the behaviour of the structural relaxation with the cage effect. We find a good agreement with the predictions of the MCT in the short and medium scale range, while at increasing length scales the interaction of water molecules with the protein's substrate induces deviations from the MCT behaviour, as found in previous studies. Besides at low temperatures the slow dynamics deviates from MCT due to hopping processes, similar to the bulk, as witnessed by a crossover from a fragile behaviour to a strong behaviour. We show that this deviation is evident at all length scales. Interestingly, we find that in the fragile region the confining cage decreases in radius with temperature while in the strong region it appears stable. 相似文献
45.
Hydrophobic polysulfone UF membranes were modified with UV irradiation and hydrophilicity increasing agents. The modifications were tested with 0.5% whey-protein solution and 0.05% lysozyme solution at pH 6 and with 0.05% bovine serum albumin solution at various pH values. UV irradiation increased flux and the hydrophilicity of the membranes. The flux increases obtained varied with pH and modification agents used and could be more than 400% compared to unmodified conditions without any loss in retention. The best retentions were obtained at pH values, where both the protein and the membrane had the same charge, and a strong electrostatic repulsion was obtained. The pores enlarged to fixed sizes, which depended on the sizes of the proteins and the range of double layer forces between proteins and membranes at different states of charge density. 相似文献
46.
Refolding human lysozyme produced as an inclusion body by urea concentration and pH gradient ion exchange chromatography 总被引:6,自引:0,他引:6
Summary A new dual-gradient ion exchange chromatographic method was developed to improve the refolding yield of human lysozyme produced
inEscherichia coli as an inclusion body. The dissolved and stretched polypeptide chain in a concentrated non-ionic denaturant was adsorbed onto
an ion exchange column and induced to refold by gradually decreasing the denaturant concentration and increasing pH in the
flowing buffer. The dual gradients of denaturant concentration and pH provided a gradual change of the solution environment
along the chromatographic column for the protein to refold, resulting in enhanced activity yield and purity. A post-separation
was also studied using size-exclusion chromatography to remove protein aggregates and mis-folded proteins after the refolding
step. 相似文献
47.
Densities and heat capacities of lysozyme in Na-acetate buffer (pH 4.2) containing 0.64 m sodium chloride at 25°C were determined
by Anton Paar 60/602 digital densimeter and differential scanning adiabatic calorimeter DASM-4 in the range of lysozyme concentration
0.000499–0.002450 m. The measurements were made after 1, 24 and 48 h of the dissolution of lysozyme in the buffer. The changes
of the values of apparent molar volumes and heat capacities in time were observed.
This revised version was published online in July 2006 with corrections to the Cover Date. 相似文献
48.
Hai Liang CUI Yong YU* W an Chun CHEN Qi KANG National Microgravity Lab Institute of Mechanics Chinese Academy of Science Beijing Institute of Physics Chinese Academy of Science Beijing 《中国化学快报》2006,17(1):101-104
Space environment is regarded as the perfect environment for the production of higher quality protein crystals since the sedimentation movement and convective flow due to the gravity is negligible under the microgravity condition of space environment. A n… 相似文献
49.
通过缩水甘油(GLY)与阿霉素(DOX)发生开环反应,赋予阿霉素活性羟基基团,以氨基己酸(ACA)为桥梁,通过共价键合方法将缩水甘油化阿霉素(GLY-DOX,GDOX)偶联于氨基己酸化纳米钻石(ND-ACA)载体上,获得具有溶菌酶响应特性的ND-ACA-GLY-DOX(NAGD)药物输送体系。 采用荧光光谱法测定了ND表面偶联ACA和DOX的量分别为(185±10.0) μg/mg和(115±5.2) μg/mg。 体外释药发现NAGD在生理环境(pH=7.4)中药物释放量很低,而在肿瘤细胞溶酶体环境中(溶菌酶存在下),酯键水解断裂,大量释放DOX。 以肝癌细胞HepG2为模型,采用细胞形态测试法表明NAGD可有效杀死肿瘤细胞。 上述研究结果表明,NAGD可作为一种良好的药物输送体系,并为共价偶联药物开拓新思维。 相似文献
50.