Abstract:
Retinol is a lipid-soluble form of vitamin A that is crucial for human visual and immune functions. The production of retinol through microbial fermentation has been the focus of recent exploration. However, the obtained titer remains limited and the product is often a mixture of retinal, retinol, and retinoic acid, necessitating purification. To achieve efficient biosynthesis of retinol in Yarrowia lipolytica, we improved the metabolic flux of β-carotene to provide sufficient precursors for retinol in this study. Coupled with the optimization of the expression level of β-carotene 15,15\'-dioxygenase, de novo production of retinol was achieved. Furthermore, Tween 80 was used as an extractant and butylated hydroxytoluene as an antioxidant to extract intracellular retinol and prevent retinol oxidation, respectively. This strategy significantly increased the level of retinol production. By optimizing the enzymes converting retinal to retinol, the proportion of extracellular retinol in the produced retinoids reached 100%, totaling 1042.3 mg/L. Finally, total retinol production reached 5.4 g/L through fed-batch fermentation in a 5 L bioreactor, comprising 4.2 g/L extracellular retinol and 1.2 g/L intracellular retinol. This achievement represents the highest reported titer so far and advances the industrial production of retinol.
摘要:
视黄醇是一种脂溶性维生素A,对人体视觉和免疫功能至关重要。通过微生物发酵生产视黄醇一直是最近探索的重点。然而,获得的滴度仍然有限,产品通常是视网膜的混合物,视黄醇,和视黄酸,需要净化。为实现解脂耶氏酵母中视黄醇的高效生物合成,在这项研究中,我们改善了β-胡萝卜素的代谢通量,为视黄醇提供了足够的前体。结合β-胡萝卜素15,15'-双加氧酶表达水平的优化,实现了视黄醇的从头生产。此外,吐温80作为萃取剂和丁基羟基甲苯作为抗氧化剂,以提取细胞内的视黄醇和防止视黄醇氧化,分别。该策略显著提高了视黄醇的生产水平。通过优化将视黄醛转化为视黄醇的酶,生产的类视黄醇中细胞外视黄醇的比例达到100%,总计1042.3mg/L最后,在5升生物反应器中通过补料分批发酵,总视黄醇产量达到5.4克/升,包含4.2g/L细胞外视黄醇和1.2g/L细胞内视黄醇。这一成就代表了迄今为止报道的最高滴度,并推进了视黄醇的工业生产。