The influence of operational parameters on lysozyme refolding using size-exclusion chromatography
The influence of operational parameters on lysozyme refolding using size-exclusion chromatography
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DOI：
10.1007/s00449-001-0271-8
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发表时间：
2001-11-01
影响因子：
3.8
通讯作者：
Chaudhuri, JB
中科院分区：
文献类型：
--
作者：
Batas, B;Chaudhuri, JB
研究了几个参数对起始浓度为 40 mg/ml 的鸡蛋清溶菌酶凝胶过滤重折叠的影响。发现重折叠不受 30 至 50°C 温度的影响，比活性 100% 恢复。在 10°C 时，观察到重折叠率降低了 20%。使用基于丙烯酰胺 (Sephacryl S100) 和葡聚糖 (Superdex 75) 的凝胶介质成功地进行了重折叠。在溶菌酶的等电点 pH 值下，柱法中的聚集受到抑制，而在基于稀释的重折叠过程中形成了蛋白质聚集体。流动相中添加了多种化合物（羧甲基纤维素、葡聚糖、蔗糖）以降低样品和流动相之间的相对粘度。只有高达 20%（重量）的蔗糖被发现不会干扰溶菌酶的重折叠。
The influence of several parameters on the gel filtration refolding of hen egg white lysozyme from a starting concentration of 40 mg/ml was investigated. Refolding was found to be unaffected by temperature between 30 and 50degreesC, giving 100% recovered specific activity. At 10degreesC a 20% reduction in refolding yield was observed. Refolding was carried out successfully with both acrylamide (Sephacryl S100)- and dextran (Superdex 75)based gel media. At the isoelectric pH of lysozyme, aggregation was suppressed in the column method, whereas protein aggregates were formed during dilution-based refolding. A number of compounds (carboxymethyl cellulose, dextran, sucrose) were added to the mobile phase to reduce the relative viscosity between the sample and mobile phase. Only sucrose, up to 20% (wt), was found not to interfere with lysozyme refolding.