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首页> 外文期刊>Protein Expression and Purification >Expression of a soluble and activatable form of bovine procarboxypeptidase A in Escherichia coli
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Expression of a soluble and activatable form of bovine procarboxypeptidase A in Escherichia coli

机译：可溶和可活化形式的牛羧肽酶A在大肠杆菌中的表达

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Bovine pancreatic procarboxypeptidase A has been overexpressed in a soluble and activatable form in Escherichia coli. When the protein was expressed under the control of bacteriophage T7 promoter in E coli ADA494 (a thioredoxin reductase deficient bacteria), a thioredoxin fusion protein was produced at relatively high level in the cytoplasm (4 mg/L culture medium). Although the recombinant protein essentially accumulated as inclusion bodies, as much as 30% of the fusion protein was recovered in a soluble form at low growth temperature and could therefore be purified to homogeneity in a single-step procedure by metal-affinity chromatography. The recombinant precursor form of bovine carboxypeptidase A was recognized by a monoclonal antibody directed against purified bovine pancreatic carboxypeptidase A. Moreover, upon tryptic activation it gave rise to an enzyme, the N-terminal sequence, molecular size,and specific activity of which were comparable to those of the enzyme derived from the native precursor purified from bovine pancreas. (C) 2002 Elsevier Science (USA). All rights reserved. [References: 31]
机译：牛胰原羧肽酶A在大肠杆菌中以可溶和可活化的形式过表达。当在大肠杆菌ADA494（一种硫氧还蛋白还原酶缺陷型细菌）的噬菌体T7启动子的控制下表达该蛋白时，会在细胞质中以相对较高的水平产生硫氧还蛋白融合蛋白（4 mg / L培养基）。尽管重组蛋白本质上是作为包涵体积累的，但高达30％的融合蛋白在低生长温度下以可溶形式被回收，因此可以通过金属亲和色谱法在一步法中纯化至均一。牛羧肽酶A的重组前体形式被针对纯化牛胰羧肽酶A的单克隆抗体识别。此外，在胰蛋白酶活化后，它产生了一种酶，其N端序列，分子大小和比活性可比来源于从牛胰腺纯化的天然前体的酶。 （C）2002 Elsevier Science（美国）。版权所有。 [参考：31]

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