Efficient secretory expression of Bacillus stearothermophilus α/β-cyclodextrin glycosyltransferase in Bacillus subtilis
Efficient secretory expression of Bacillus stearothermophilus α/β-cyclodextrin glycosyltransferase in Bacillus subtilis
复制标题
枯草芽孢杆菌中嗜热脂肪芽孢杆菌α/β-环糊精糖基转移酶的高效分泌表达
DOI：
10.1016/j.jbiotec.2021.03.011
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发表时间：
2021-03-21
影响因子：
4.1
通讯作者：
Wu, Jing
中科院分区：
文献类型：
--
作者：
Su, Lingqia;Li, Yunfei;Wu, Jing
嗜热脂肪芽孢杆菌？/？-环糊精糖基转移酶（β/β- CGTase）是一种优良的转糖基酶，具有广泛的食品应用前景，但在枯草芽孢杆菌中表达量较低。在本研究中，最佳的信号肽？/？-从B中的173个信号肽中筛选CGTase表达。subtilis WS 11. The？/？-在3L发酵罐中酶活达到151.93U. mL-1，但产生了大量的包涵体。N端12个氨基酸残基的氨基酸序列CGTase然后被替换为？-来自同一家族的CGTase，具有高百分比的促进紊乱的氨基酸。结果表明，包涵体明显减少，酶活提高到249.35 U mL-1，是原菌株的2.3倍。最后，对B.将与脂肽生物表面活性剂合成相关的枯草芽孢杆菌WS 11基因敲除，以产生B。subtilis WS 13.当B.以枯草芽孢杆菌WS 13为出发菌株，CGTase在3-L发酵罐中的发酵比B需要的发酵剂少70%。subtilis WS 11.此外，WS 13的产酶和生长与WS 11相当。该研究对今后高效放大生产？/？-？具有重要意义CGTase。
Bacillus stearothermophilus ?/?-cyclodextrin glycosyltransferase (?/?-CGTase) is an excellent transglycosylase with broad potential for food application, but its expression level is low in Bacillus subtilis. In this study, the optimal signal peptide for ?/?-CGTase expression was screened from 173 signal peptides in B. subtilis WS11. The ?/?-CGTase activity in a 3-L fermentor reached 151.93 U. mL-1, but substantial amounts of inclusion bodies were produced. The N-terminal 12 amino acids of ?/?-CGTase were then replaced with the N-terminal 15 amino acids of a ?-CGTase from the same family that has a high percentage of disorder-promoting amino acids. As a result, the inclusion bodies were significantly reduced, and the enzyme activity increased to 249.35 U mL-1, 2.3 times that of the strain constructed previously. Finally, the ppsE and sfp genes of B. subtilis WS11, which are related to lipopeptide biosurfactant synthesis, were knocked out to produce B. subtilis WS13. When B. subtilis WS13 was used to produce ?/?-CGTase in a 3-L fermentor, 70 % less defoaming agent was required than with B. subtilis WS11. Furthermore, enzyme production and growth of WS13 were equivalent to those of WS11. This study is of great significance for future research to efficiently scale-up production of ?/?-CGTase.