Amylosucrase (EC 2.4.1.4) is a versatile enzyme with significant potential in biotechnology and food production. To facilitate its efficient preparation, a novel expression strategy was implemented in Bacillus licheniformis for the secretory expression of Neisseria polysaccharea amylosucrase (NpAS). The host strain B. licheniformis CBBD302 underwent genetic modification through the deletion of sacB, a gene responsible for encoding levansucrase that synthesizes extracellular levan from sucrose, resulting in a levan-deficient strain, B. licheniformis CBBD302B. NpAS was successfully expressed in B. licheniformis CBBD302B using the highly efficient Sec-type signal peptide SamyL, but its extracellular translocation was unsuccessful. Consequently, the expression of NpAS via the twin-arginine translocation (TAT) pathway was investigated using the signal peptide SglmU. The study revealed that NpAS could be effectively translocated extracellularly through the TAT pathway, with the signal peptide SglmU facilitating the process. Remarkably, 62.81% of the total expressed activity was detected in the medium. This study marks the first successful secretory expression of NpAS in Bacillus species host cells, establishing a foundation for its future efficient production.

中文翻译：

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地衣芽孢杆菌通过双精氨酸易位途径分泌表达淀粉蔗糖酶

淀粉蔗糖酶 (EC 2.4.1.4) 是一种多功能酶，在生物技术和食品生产中具有巨大潜力。为了促进其高效制备，在地衣芽孢杆菌中实施了一种新的表达策略，用于分泌表达多糖奈瑟菌淀粉蔗糖酶（NpAS）。宿主菌株地衣芽孢杆菌 CBBD302 通过删除 sacB（负责编码从蔗糖合成胞外果聚糖的果聚糖蔗糖酶的基因）进行遗传修饰，产生了左聚糖缺陷菌株，即地衣芽孢杆菌 CBBD302B。利用高效的Sec型信号肽SamyL在地衣芽孢杆菌CBBD302B中成功表达NpAS，但其胞外转位不成功。因此，使用信号肽 SglmU 研究了 NpAS 通过双精氨酸易位 (TAT) 途径的表达。研究表明，NpAS 可以通过 TAT 途径有效地转移到细胞外，信号肽 SglmU 可以促进这一过程。值得注意的是，在培养基中检测到了总表达活性的 62.81%。该研究标志着NpAS首次在芽孢杆菌属宿主细胞中成功分泌表达，为其未来高效生产奠定了基础。