Braia Mauricio, Ferrero Maximiliano, Rocha María Victoria, Loureiro Dana, Tubio Gisela, Romanini Diana
Laboratory of Physical Chemistry Applied to Bioseparation-Rosario, Faculty of Biochemical and Pharmaceutical Sciences, National University of Rosario, Suipacha 531, S2002RLK Rosario, Argentina.
Protein Expr Purif. 2013 Sep;91(1):91-5. doi: 10.1016/j.pep.2013.07.008. Epub 2013 Jul 23.
The formation of insoluble complexes between enzymes and polyelectrolytes is a suitable technique for isolating these biomolecules from natural sources, because it is a simple and rapid technique that allows the concentration of the protein. This technique can be used in most purification protocols at the beginning of the downstream process. The aim of this investigation is to isolate papain from Carica papaya latex by precipitation of insoluble complexes between this enzyme and poly (vinyl sulfonate). The papain-poly (vinyl sulfonate) complex was insoluble at pH lower than 6, with a PVS/PAP stoichiometric ratio of 1:279. Ionic strength affected the complex formation. The presence of the polymer increased the enzymatic activity and protected the enzyme from autodegradation. The optimal conditions for the formation of insoluble papain-polyelectrolyte complex formation were applied to C. papaya latex and a high recovery was obtained (around 86%) and a purification factor around 2. This method can be applied as an isolation method of papain from C. papaya latex or as a first step in a larger purification strategy.
酶与聚电解质之间形成不溶性复合物是从天然来源分离这些生物分子的一种合适技术,因为它是一种简单快速的技术,能够实现蛋白质的浓缩。该技术可用于大多数纯化方案的下游处理初期。本研究的目的是通过沉淀木瓜蛋白酶与聚(乙烯基磺酸盐)之间的不溶性复合物,从番木瓜乳胶中分离木瓜蛋白酶。木瓜蛋白酶 - 聚(乙烯基磺酸盐)复合物在pH低于6时不溶,PVS/PAP化学计量比为1:279。离子强度影响复合物的形成。聚合物的存在提高了酶活性并保护酶免于自降解。将形成不溶性木瓜蛋白酶 - 聚电解质复合物的最佳条件应用于番木瓜乳胶,获得了较高的回收率(约86%)和纯化因子约为2。该方法可作为从番木瓜乳胶中分离木瓜蛋白酶的方法,或作为更大规模纯化策略的第一步。