Selecting Endogenous Promoters for Improving Biosynthesis of Squalene in Schizochytrium sp.

Abstract

Squalene (C30H50) is an acyclic triterpenoid compound renowned for its myriad physiological functions, such as anticancer and antioxidative properties, rendering it invaluable in both the food and pharmaceutical sectors. Due to the natural resource constraints, microbial fermentation has emerged as a prominent trend. Schizochytrium sp., known to harbor the intact mevalonate acid (MVA) pathway, possesses the inherent capability to biosynthesize squalene. However, there is a dearth of reported key genes in both the MVA and the squalene synthesis pathways, along with the associated promoter elements for their modification. This study commenced by cloning and characterizing 13 endogenous promoters derived from transcriptome sequencing data. Subsequently, five promoters exhibiting varying expression intensities were chosen from the aforementioned pool to facilitate the overexpression of the squalene synthase gene squalene synthetase (SQS), pivotal in the MVA pathway. Ultimately, a transformed strain designated as SQS-3626, exhibiting squalene production 2.8 times greater than that of the wild-type strain, was identified. Finally, the optimization of nitrogen source concentrations and trace element contents in the fermentation medium was conducted. Following 120 h of fed-batch fermentation, the accumulated final squalene yield in the transformed strain SQS-3626 reached 2.2g/L.