Enhanced Production of Transglutaminase in Streptomyces mobaraensis through Random Mutagenesis and Site-Directed Genetic Modification
Enhanced Production of Transglutaminase in Streptomyces mobaraensis through Random Mutagenesis and Site-Directed Genetic Modification
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通过随机诱变和定点遗传修饰增强茂原链霉菌中转谷氨酰胺酶的产量。
DOI：
10.1021/acs.jafc.1c00645
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发表时间：
2021-03-02
影响因子：
6.1
通讯作者：
Liu, Song
中科院分区：
文献类型：
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作者：
Yin, Xiaoqiang;Li, Yangyang;Liu, Song
链霉菌转谷氨酰胺酶 (TGase) 广泛用于改善食品质地特性。在本研究中，采用随机诱变和定点遗传修饰来提高茂原链霉菌的 TGase 产量。首先，对S. mobaraensis DSM40587（smWT）进行常压和室温等离子体诱变，然后从约3000×25（圆形）突变体中筛选出TGase产量增加5.5倍的突变体（smY2019）。与 smWT 相比，smY2019 的 TGase mRNA 水平高出 3.2 倍，并且 TGase 启动子的 -10 区域内有两个位点突变。 TGase 缺陷型 S. mobaraensis 中的重组表达分析表明，突变的 TGase 启动子比野生型启动子更强大。最后，我们将两个额外的TGase表达盒整合到smY2019基因组中，产生重组菌株smY2019-3C，与smY2019相比，TGase产量增加了103%。 smY2019-3C 菌株的 TGase 产量为 40 U/mL，可能是工业化生产 TGase 的合适候选菌株。
Streptomyces transglutaminase (TGase) is widely used to improve food texture properties. In this study, random mutagenesis and site-directed genetic modification were used to improve the production of TGase in Streptomyces mobaraensis. First, S. mobaraensis DSM40587 (smWT) was subjected to atmospheric and room-temperature plasma mutagenesis, and then a mutant (smY2019) with a 5.5-fold increase in TGase yield was screened from approximately 3000 × 25 (round) mutants. Compared to smWT, smY2019 exhibits a 3.2-fold higher TGase mRNA level and two site mutations within the -10 region of the TGase promoter. The recombinant expression analysis in the TGase-deficient S. mobaraensis suggests that the mutated TGase promoter is more robust than the wild-type one. Finally, we integrated two additional TGase expression cassettes into the smY2019 genome, yielding the recombinant strain smY2019-3C with a 103% increase in TGase production compared to smY2019. The smY2019-3C strain with 40 U/mL of TGase yield could be a suitable candidate for the industrial production of TGase.