A coordinated framework for enhancing oxygen dynamics to improve the production of sesquiterpene

Abstract

Microbial production of sesquiterpene like β-farnesene is challenging owing to high energy demands, redox imbalances, and oxygen supply constraints. However, the underlying mechanisms governing these limitations remain poorly understood, making it challenging to optimize production through traditional metabolic engineering strategies. Here, we show a multi-scale framework focused on oxygen dynamics by integrating bioreactor optimization, transcriptomic sequencing, and metabolic engineering to efficiently produce β-farnesene. Starting with improved oxygen supply, we demonstrated high oxygen demand during β-farnesene synthesis and alleviated limitations through an impeller design, which enhanced the oxygen transfer efficiency. Modifications of genes GRE2, SOD2, ERG2, and ALP1 boosted the production capacity, while the expression of hemoglobin improved oxygen utilization and redox homeostasis. These strategies led to a 242% enhancement in β-farnesene production, accompanied by substantial reductions in glucose and oxygen consumption. This work presents a comprehensive strategy to enhance sesquiterpene biosynthesis and provides insights into overcoming oxygen constraints in microbial cell factories.