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首页> 外文期刊>Separation and Purification Technology >Studies of protein partition in non conventional aqueous two-phase systems as method to purify trypsinogen and alpha-chymotrypsinogen from bovine pancreas
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Studies of protein partition in non conventional aqueous two-phase systems as method to purify trypsinogen and alpha-chymotrypsinogen from bovine pancreas

机译：研究非常规水两相系统中的蛋白质分配作为从牛胰腺中纯化胰蛋白酶原和α-胰凝乳蛋白酶原的方法

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摘要

Aqueous two phase systems (ATPSs) formed by polyethyleneglycol (PEG) and sodium tartrate (NaTart) pH 5.00 are non conventional systems of low impact. Their ability to purify trypsinogen (TRPz) and alpha-chymotrypsinogen (ChTRPz) from bovine pancreas was evaluated. The effect of several variables such as molecular weight of PEG, tie line length and phase-volume ratio on zymogen partitioning was explored. Those systems formed by PEG of molecular weight 600 (20.62%, w/w) and NaTart (12.85%, w/w) with a phase-volume ratio equal to one were found to evidence the best purification performance, and were employed to design a two-stage purification process. Firstly, a ATPS extraction step yields both to recover most of pancreatic TRPz and ChTRPz (76% and 87% respectively) in the PEG600 enriched-top phase and to eliminate other pancreatic homogenate components - cell debris, nucleic acids and lipids - which either partition into the bottom phase or precipitate at the interface. Then, the top phase - from this first extraction step - is mixed with a given quantity of PEG6000 in order to induce the formation of a new aqueous two phase system in which the back extraction of both zymogens takes place. This second stage allows to obtain a mixture of both zymogens highly pure (approx =95%) in a bottom phase with low PEG concentration (3.70%, w/w) and therefore suitable for being used in multiple applications.
机译：由聚乙二醇（PEG）和酒石酸钠（NaTart）pH 5.00形成的水两相系统（ATPS）是非传统的低冲击系统。评价了它们从牛胰腺中纯化胰蛋白酶原（TRPz）和α-胰凝乳蛋白酶原（ChTRPz）的能力。探索了PEG分子量，束线长度和相体积比等变量对酶原分配的影响。发现那些由分子量为600（20.62％，w / w）的PEG和NaTart（12.85％，w / w）的相-体积比等于1的体系形成的系统具有最佳的纯化性能，并被用于设计两阶段纯化过程。首先，ATPS提取步骤既可回收富含PEG600的上层液相中的大部分胰TRPz和ChTRPz（分别为76％和87％），又可消除其他胰匀浆成分（细胞碎片，核酸和脂质）进入底部相或在界面处沉淀。然后，将来自第一萃取步骤的上层相与给定量的PEG6000混合，以诱导形成新的含水两相系统，在其中发生两种酶原的反萃取。该第二阶段允许在底部相中获得高纯度（大约= 95％）的两种酶原的混合物，而PEG浓度低（3.70％，w / w），因此适合用于多种应用。

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