制备了一种能有效分离纯化木瓜蛋白酶的金属螯合膜.该膜以包裹壳聚糖的尼龙膜为基质,通过环氧氯丙烷交联活化、亚氨基二乙酸偶联、金属镍离子螯合制备得到.研究了吸附时间、溶液的pH、溶液的离子强度等因素对吸附的影响,考察了吸附模型,并对其色谱性能和再生性进行探讨.结果表明,最佳吸附条件pH 10.0,反应时间60 min,离子强度0 mol/L,吸附量达67.2 mg/g,吸附行为满足Freundlieh等温吸附模型,其色谱过程中的洗脱剂为1.0 mol/L NaSCN的Tris-HCI(pH5.0)其纯化倍数可以达到19.1倍.吸附洗脱实验重复3次,吸附率仪降低8.7%,表明该模具有良好的稳定性,重复性和再生性,能有效用于木瓜蛋白酶的分离.
参考文献
| [1] | Ghosh S .Conformational study of papain in the presence of sodium dodecyl sulfate in aqueous medium[J].Colloids and Surfaces, B. Biointerfaces,2005(2/3):209-216. |
| [2] | Sangeetha K;Abraham TE .Chemical modification of papain for use in alkaline medium[J].Journal of molecular catalysis, B. Enzymatic,2006(3/6):171-177. |
| [3] | Szabo A;Kotorman M;Laczko I;Simon LM .Spectroscopic studies of stability of papain in aqueous organic solvents[J].Journal of molecular catalysis, B. Enzymatic,2006(1/2):43-48. |
| [4] | Sarote Nitsawang;Rajni Hatti-Kaul;Pawinee Kanasawud .Purification of papain from Carica papaya latex:Aqueous two-phase extraction versus two-step salt precipitation[J].Enzyme and Microbial Technology,2006(5):1103-1107. |
| [5] | Tomhaceini D;Mocali A;Weber E et al.A cystatinbased affinity procedure for the isolation and analysis of papain-like eysteine proteinases from tissue extracts[J].Analytical Biochemistry,2001,289:231-238. |
| [6] | Govrin E;Levine A .Purification of active cysteine proteases by affinity chromatography with attached E-64 inhibitor.[J].Protein Expression and Purification,1999(3):247-250. |
| [7] | Hu HL;Wang MY;Chung CH;Suen SY .Purification of VP3 protein of infectious bursal disease virus using nickel ion-immobilized regenerated cellulose-based membranes[J].Journal of chromatography, B. Analytical technologies in the biomedical and life sciences,2006(2):76-84. |
| [8] | Nie HL;Zhu LM .Adsorption of papain with Cibacron Blue F3GA carrying chitosan-coated nylon affinity membranes[J].International Journal of Biological Macromolecules: Structure, Function and Interactions,2007(3):261-267. |
| [9] | Han Lei;Wei Wang;Ling-Li Chen;Xiao-Cong Li;Bin Yi;Le Deng .The preparation and catalytically active characterization of papain immobilized on magnetic composite microspheres[J].Enzyme and Microbial Technology,2004(1):15-21. |
| [10] | 陈钧辉.生物化学实验[M].北京:科学出版社,2003:63. |
| [11] | Arica M Y;Testereci H N;Denizli Adil .Dye-ligand and metal chelate poly(2-hydroxyethylmethacrylate)membranes for affinity separation of proteins[J].J Chrornatogr A,1998,799:83-91. |
| [12] | Bayramoglu G;Celik G;Arica MY .Immunoglobulin G adsorption behavior of L-histidine ligand attached and Lewis metal ions chelated affinity membranes[J].Colloids and Surfaces, A. Physicochemical and Engineering Aspects,2006(1/3):75-85. |
上一张
下一张
上一张
下一张
计量
- 下载量()
- 访问量()
文章评分
- 您的评分：
-
10%
-
20%
-
30%
-
40%
-
50%