Systematic Screening of Optimal Signal Peptides for Secretory Production of Heterologous Proteins in Bacillus subtilis,Journal of Agricultural and Food Chemistry

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Systematic Screening of Optimal Signal Peptides for Secretory Production of Heterologous Proteins in Bacillus subtilis
Journal of Agricultural and Food Chemistry ( IF 5.7 ) Pub Date : 2018-11-22 00:00:00 , DOI: 10.1021/acs.jafc.8b04183
Gang Fu _{1,

2} , Jinlan Liu _{1,

2} , Jinshan Li _{1,

2} , Beiwei Zhu ₃ , Dawei Zhang _{1,

2,

3,

4}

Affiliation

Bacillus subtilis is widely used for large-scale industrial production of heterologous proteins. Because of its high intrinsic secretory capacity, it can efficiently secrete proteins into the culture supernatant via the general Sec-type secretion pathway. In this study, the α-amylase AmyS was used as a reporter to construct a library encompassing 173 Sec-type signal peptides (SPs) from B. subtilis using a fast, sequence-independent method. The resulting library DNA which harbored different signal peptides in the expression vector was used to transform B. subtilis directly at high efficiency, and 15 SPs which produced a significantly increased yield of AmyS were identified using a starch-iodine-based high-throughput assay. Furthermore, the correlation between the sequences of the best-performing signal peptides and their secretion efficiency was analyzed, which revealed several common properties of these SPs. Finally, high-cell-density fermentation of the α-amylase-producing strain with the best-performing signal peptide yielded a maximum of 5086 U/mL amylase at 66 h with a high productivity of 77.1 U/mL·h.

中文翻译：

枯草芽孢杆菌分泌异源蛋白的最佳信号肽的系统筛选

枯草芽孢杆菌广泛用于异源蛋白质的大规模工业生产。由于其固有的高分泌能力，它可以通过一般的Sec型分泌途径将蛋白质有效地分泌到培养上清液中。在这项研究中，使用α-淀粉酶AmyS作为报告基因，以快速，独立于序列的方法构建了一个库，该库包含来自枯草芽孢杆菌的173个Sec型信号肽（SP）。在表达载体中带有不同信号肽的所得文库DNA被用于转化枯草芽孢杆菌。使用基于淀粉-碘的高通量检测方法鉴定出15个SPs，它们可以显着提高AmyS的产量，并且效率很高。此外，分析了最佳信号肽序列与其分泌效率之间的相关性，揭示了这些SP的几个共同特性。最后，具有最佳信号肽的α-淀粉酶生产菌株的高细胞密度发酵在66 h时产生的最大淀粉酶为5086 U / mL，效率为77.1 U / mL·h。

更新日期：2018-11-22

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