BACKGROUND Recombinant proteins are often engineered with an N-terminal signal peptide, which facilitates their secretion to the oxidising environment of the periplasm (gram-negative bacteria) or the culture supernatant (gram-positive bacteria). A commonly encountered problem is that the signal peptide influences the synthesis and secretion of the recombinant protein in an unpredictable manner. A molecular understanding of this phenomenon is highly sought after, as it could lead to improved methods for producing recombinant proteins in bacterial cell factories. RESULTS Herein we demonstrate that signal peptides contribute to an unpredictable translation initiation region. A directed evolution approach that selects a new translation initiation region, whilst leaving the amino acid sequence of the signal peptide unchanged, can increase production levels of secreted recombinant proteins. The approach can increase production of single chain antibody fragments, hormones and other recombinant proteins in the periplasm of E. coli. CONCLUSIONS The study demonstrates that signal peptide performance is coupled to the efficiency of the translation initiation region.

中文翻译：

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通过翻译起始区的定向进化，可增加大肠杆菌中周质蛋白的产量。

背景技术重组蛋白通常用N端信号肽进行工程改造，这有助于它们分泌到周质（革兰氏阴性细菌）或培养上清液（革兰氏阳性细菌）的氧化环境中。普遍遇到的问题是信号肽以不可预测的方式影响重组蛋白的合成和分泌。人们强烈要求对这种现象进行分子理解，因为这可能导致改进的在细菌细胞工厂中生产重组蛋白的方法。结果在本文中我们证明信号肽有助于不可预测的翻译起始区。一种定向进化方法，它选择一个新的翻译起始区，同时使信号肽的氨基酸序列保持不变，可以增加分泌的重组蛋白的生产水平。该方法可以增加大肠杆菌周质中单链抗体片段，激素和其他重组蛋白的产量。结论该研究表明信号肽的性能与翻译起始区的效率有关。