过表达Yap1增强黑曲霉葡萄糖氧化酶分泌

Enhanced Glucose Oxidase Production in Aspergillus niger by Yap1 Overexpression

摘要:

过量活性氧积累会引起氧化应激，破坏胞内稳态。该研究进行了黑曲霉野生型、转录因子敲除株ΔYap1在正常和氧化应激条件下的全基因组规模的转录差异研究。结果表明：当黑曲霉遭遇氧化应激时，Yap1通过激活超氧化物歧化酶、过氧化物酶等抗氧化酶的表达，增强其抗活性氧的能力。同时构建了葡萄糖氧化酶表达菌株，并将转录因子Yap1整合至此菌株基因组中，该菌株胞内活性氧水平较出发菌株降低了55%，达到了283.61 A.U./mg，并且葡萄糖氧化酶的酶活提高了102%，达到了321.36 U/mL，总分泌蛋白水平提升了38%，达到了26.11 mg/mL。该研究通过转录组测序揭示了黑曲霉是以转录因子Yap1为主应对胞内氧化应激，并且通过过表达Yap1可以降低其胞内活性氧水平，同时也增加其蛋白分泌能力，为工业提高蛋白（酶）的产量提供参考。

Abstract:

Excessive accumulation of reactive oxygen species (ROS) can lead to oxidative stress, thus disrupting intracellular homeostasis. A genome-wide transcriptional analysis was performed to compare the Aspergillus niger wild type and the transcription factor knockout strain ΔYap1 under normal and oxidative stress conditions. The results demonstrated that, in response to oxidative stress, Yap1 in A. niger activated the expression of antioxidant enzymes, such as superoxide dismutase and peroxidase, which enhanced its resistance to ROS. Additionally, a glucose oxidase-expressing strain was generated, and the transcription factor Yap1 was integrated into the genome of this strain. This process resulted in a 55% reduction in the intracellular ROS level, which reached 283.61 A.U./mg in the modified strain compared with that in the parent strain. Furthermore, the glucose oxidase activity increased by 102%, reaching 321.36 U/mL, and the total protein secretion increased by 38%, reaching 26.11 mg/mL. Transcriptome sequencing revealed that A. niger predominantly utilizes Yap1 to cope with intracellular oxidative stress. Through Yap1 overexpression, the intracellular ROS level was reduced and the protein secretion capacity was enhanced. Overall, these findings provide valuable insights for increasing industrial protein (enzyme) production.