Abstract：
7-Dehydrocholesterol (7-DHC) is widely present in various organisms and is an important precursor of vitamin D3. Despite significant improvements in the biosynthesis of 7-DHC, it remains insufficient to meet the industrial demands. In this study, we reported high-level production of 7-DHC in an industrial Saccharomyces cerevisiae leveraging subcellular organelles. Initially, the copy numbers of DHCR24 were increased in combination with sterol transcriptional factor engineering and rebalanced the redox power of the strain. Subsequently, the effects of compartmentalizing the post-squalene pathway in peroxisomes were validated by assembling various pathway modules in this organelle. Furthermore, several peroxisomes engineering was conducted to enhance the production of 7-DHC. Utilizing the peroxisome as a vessel for partial post-squalene pathways, the potential of yeast for 7-dehydrocholesterol production was demonstrated by achieving a 26-fold increase over the initial production level. 7-DHC titer reached 640.77 mg/L in shake flasks and 4.28 g/L in a 10 L bench-top fermentor, the highest titer ever reported. The present work lays solid foundation for large-scale and cost-effective production of 7-DHC for practical applications.
摘要：
7-脱氢胆固醇(7-DHC)广泛存在于各种生物体中,是维生素D3的重要前体。尽管7-DHC的生物合成有显著改善，它仍然不足以满足工业需求。在这项研究中,我们报道了利用亚细胞器的工业酿酒酵母中7-DHC的高水平生产。最初，DHCR24的拷贝数与固醇转录因子工程结合增加，并重新平衡了菌株的氧化还原能力。随后，通过在该细胞器中组装各种途径模块，验证了在过氧化物酶体中分隔后角鲨烯途径的作用。此外，进行了几种过氧化物酶体工程以提高7-DHC的产量。利用过氧化物酶体作为部分后角鲨烯途径的容器，酵母生产7-脱氢胆固醇的潜力通过比初始生产水平增加26倍来证明.7-DHC滴度在摇瓶中达到640.77mg/L，在10L台式发酵罐中达到4.28g/L，有史以来的最高滴度。本工作为7-DHC在实际应用中的大规模和成本有效的生产奠定了坚实的基础。
