桦木酸（BA）是从羽扇豆醇中提取的羽扇烷型五环三萜类天然产物，具有良好的抗炎和抗肿瘤活性。目前，BA主要通过植物提取生产，这极大地限制了其广泛使用。在本研究中，我们研究了酿酒酵母中BA的从头合成，为了促进疏水性BA的合成和储存，我们采用了过氧化物酶体和脂滴的双工程策略来构建BA生物合成途径。通过表达白桦来源的羽扇豆醇 C-28 氧化酶 (BPLO) 和拟南芥来源的 ATR1，我们成功开发了 BA 生产菌株，并对 BPLO 和 ATR1 之间的连接子进行了多次表达优化，BA 滴度达到 77.53 mg/L随后在 5 L 生物反应器中通过补料分批发酵达到 205.74 mg/L。在这项研究中，我们开发了一种在工程酿酒酵母中从头合成 BA 及其直接前体羽扇豆醇的可行方法。版权所有 © 2024 Elsevier Inc. 保留所有权利。

Betulinic acid (BA) is a lupinane-type pentacyclic triterpenoid natural product derived from lupeol that has favorable anti-inflammatory and anti-tumor activities. Currently, BA is mainly produced via botanical extraction, which significantly limits its widespread use. In this study, we investigated the de novo synthesis of BA in Saccharomyces cerevisiae, and to facilitate the synthesis and storage of hydrophobic BA, we adopted a dual-engineering strategy involving peroxisomes and lipid droplets to construct the BA biosynthetic pathway. By expressing Betula platyphylla-derived lupeol C-28 oxidase (BPLO) and Arabidopsis-derived ATR1, we succeeded in developing a BA-producing strain and following multiple expression optimizations of the linker between BPLO and ATR1, the BA titer reached 77.53 mg/L in shake flasks and subsequently reached 205.74 mg/L via fed-batch fermentation in a 5-L bioreactor. In this study, we developed a feasible approach for the de novo synthesis of BA and its direct precursor lupeol in engineered S. cerevisiae.Copyright © 2024 Elsevier Inc. All rights reserved.