Structured model to predict intracellular amino acid shortages during recombinant protein overexpression in E-coli
Structured model to predict intracellular amino acid shortages during recombinant protein overexpression in E-coli
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DOI：
10.1016/s0168-1656(01)00347-9
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发表时间：
2002-02-28
影响因子：
4.1
通讯作者：
Harcum, SW
中科院分区：
文献类型：
--
作者：
Harcum, SW
重组蛋白过表达和经典的严格反应已被证明诱导相同的蛋白酶。由于严格反应是细胞内氨基酸短缺的结果，因此假设重组蛋白的过表达也引起细胞内氨基酸短缺。建立了重组大肠杆菌的结构化非分离动力学数学模型，用于预测重组蛋白过量表达过程中细胞内氨基酸的短缺，从而诱导严格反应。两个模型重组蛋白进行了检查，氯霉素乙酰转移酶（CAT）和“平均蛋白质”。该模型预测在CAT过表达期间芳香族氨基酸短缺，如基于CAT的氨基酸含量所预测的。该模型还预测在CAT过表达期间细胞内丙氨酸家族氨基酸库的短缺。这是出乎意料的，因为与平均E. coli蛋白。该模型预测丙氨酸，戊二酸，天冬氨酸家族的氨基酸短缺重组的“平均蛋白”过表达。此外，该模型预测的核糖体池减少在诱导的重组蛋白，这与发表的实验结果一致。因此，结构化动力学模型能够预测氨基酸短缺，这可能会导致严格的反应和蛋白酶活性升高。(C)2002 Elsevier Science B.V.保留所有权利。
Recombinant protein overexpression and the classical stringent response have been shown to induce the same proteases. Since the stringent response was the result of an intracellular amino acid shortage, it was hypothesized that the overexpression of the recombinant protein also caused an intracellular amino acid shortage. A structured non-segregated kinetic mathematical model of recombinant Escherichia coli was developed to predict intracellular amino acid shortages during recombinant protein overexpression, and thus the induction of the stringent response. Two model recombinant proteins were examined, chloramphenicol acetyl-transferase (CAT) and an 'average protein'. The model predicted an aromatic amino acid shortage during CAT overexpression, as predicted based on the CAT's amino acid content. The model also predicted a shortage of the intracellular alanine family amino acid pool during CAT overexpression. This was unexpected due to the relatively low content of alanine family amino acids in CAT compared to the average E. coli protein. The model predicted alanine, glutarate, and aspartate family amino acid shortages during recombinant 'average protein' overexpression. Additionally, the model predicted a decrease in the ribosome pool at induction for both recombinant proteins, which agrees with published experimental results. Thus, the structured kinetic model was able to predict amino acid shortages, that could potentially cause a stringent response and elevated protease activity. (C) 2002 Elsevier Science B.V. All rights reserved.