Expression and antimicrobial activity of the recombinant bovine lactoferricin in Pichia pastoris,Synthetic and Systems Biotechnology

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Expression and antimicrobial activity of the recombinant bovine lactoferricin in Pichia pastoris
Synthetic and Systems Biotechnology Pub Date : 2023-12-25 , DOI: 10.1016/j.synbio.2023.12.002
Xueqin Lv _{1,

2,

3} , Yuting Zhang _{1,

2} , Lingrui Wang _{1,

2} , Shixiu Cui ₄ , Yanfeng Liu _{1,

2} , Jianghua Li _{1,

2,

4} , Guocheng Du _{1,

2} , Long Liu _{1,

2,

3,

5}

Affiliation

Lactoferricin, a multifunctional peptide located in the -terminal region of lactoferrin, has a broad-spectrum bacteriostatic activity. It is a promising candidate as a food additive and immune fortification agent and does not have the risks associated with drug residues and drug resistance. First, we performed promoter and host cell screening to achieve the recombinant expression of lactoferricin in , showing an initial titer of 19.5 mg/L in X-33 using P promoter. Second, we constructed a 0030-α hybrid signal peptide by fusing the 0030 signal peptide with the pro-sequence of α-factor secretory signal peptide. This further increased the production of lactoferricin, with a titer of 28.8 mg/L in the fermentation supernatant in the shaking flask. Next, we increased the expression of lactoferricin by fusing it with anionic antioxidant peptides. The neutralization of positive charges yielded a titer of 55.3 mg/L in the shaking flask, and a highest titer of 193.9 mg/L in a 3-L bioreactor. The antimicrobial activity analysis showed that recombinant-expressed lactoferricin exhibited potent antibacterial activity against , and . This study provides a reference for the construction of microbial cell factories capable of efficiently synthesizing antimicrobial peptides.

中文翻译：

重组牛乳铁蛋白在毕赤酵母中的表达及抗菌活性

乳铁蛋白是一种位于乳铁蛋白β-末端区域的多功能肽，具有广谱抑菌活性。它是一种很有前途的食品添加剂和免疫增强剂候选者，并且不存在与药物残留和耐药性相关的风险。首先，我们进行了启动子和宿主细胞筛选，以实现乳铁素在中的重组表达，使用 P 启动子在 X-33 中显示初始滴度为 19.5 mg/L。其次，我们通过将0030信号肽与α因子分泌信号肽的前序列融合来构建0030-α杂合信号肽。这进一步增加了乳铁素的产量，摇瓶中发酵上清液中乳铁素的效价为28.8 mg/L。接下来，我们通过将乳铁蛋白与阴离子抗氧化肽融合来增加其表达。正电荷中和在摇瓶中产生的滴度为 55.3 mg/L，在 3 L 生物反应器中的最高滴度为 193.9 mg/L。抗菌活性分析表明，重组表达的乳铁素对、和表现出有效的抗菌活性。该研究为构建高效合成抗菌肽的微生物细胞工厂提供了参考。

更新日期：2023-12-25

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