Enhancement of protein production in Aspergillus niger by engineering the antioxidant defense metabolism

Research on protein production holds significant importance in the advancement of food technology, agriculture, pharmaceuticals, and bioenergy. Aspergillus niger stands out as an ideal microbial cell factory for the production of food-grade proteins, owing to its robust protein secretion capacity and excellent safety profile. However, the extensive oxidative folding of proteins within the endoplasmic reticulum (ER) triggers ER stress, consequently leading to protein misfolding reactions. This stressful phenomenon results in the accelerated generation of reactive oxygen species (ROS), thereby inducing oxidative stress. The accumulation of ROS can adversely affect intracellular DNA, proteins, and lipids.

In this study, we enhanced the detoxification of ROS in A. niger (SH-1) by integrating multiple modules, including the NADPH regeneration engineering module, the glutaredoxin system, the GSH synthesis engineering module, and the transcription factor module. We assessed the intracellular ROS levels, growth under stress conditions, protein production levels, and intracellular GSH content. Our findings revealed that the overexpression of Glr1 in the glutaredoxin system exhibited significant efficacy across various parameters. Specifically, it reduced the intracellular ROS levels in A. niger by 50%, boosted glucoamylase enzyme activity by 243%, and increased total protein secretion by 88%.

The results indicate that moderate modulation of intracellular redox conditions can enhance overall protein output. In conclusion, we present a strategy for augmenting protein production in A. niger and propose a potential approach for optimizing microbial protein production system.