Malonate is a high-value chemical that can be used to produce value-added compounds. Due to the toxic by-products and low product yield for malonate production through hydrolysis of cyanoacetic acid, microbial production methods have attracted significant attention. Previously, the β-alanine pathway has been engineered in Escherichia coli for malonate production. In this study, the β-alanine pathway was constructed in Saccharomyces cerevisiae by introducing the heterologous genes of BcBAPAT and TcPAND to convert l-aspartate to malonic semialdehyde, combining with co-expression genes of AAT2 and UGA2 to improve precursor supply and malonate producing. Through delta sequence-based integration of the two heterologous genes, the engineered strain produced with 7.21 mg/L malonate was screened. Further, replaced the succinic semialdehyde dehydrogenase gene UGA2 with yneI from E. coli which was utilized to produce malonate in previous study, increased the malonate titer to 7.96 mg/L in flask culture. Following optimization, fermentation of the final engineered strain in shake flasks yielded a maximum malonate titer of 12.83 mg/L, and this was increased to 91.53 mg/L during fed-batch fermentation in a 5 L bioreactor which increased by two-fold compared with that of the engineered strain overexpressing UGA2.

丙二酸盐是一种高价值化学品，可用于生产增值化合物。由于氰基乙酸水解生产丙二酸的副产物有毒且产品收率低，微生物生产方法引起了人们的极大关注。此前，β-丙氨酸途径已在大肠杆菌中进行工程改造，用于生产丙二酸。本研究通过引入BcBAPAT和TcPAND异源基因将L-天冬氨酸转化为丙二酸半醛，结合AAT2和UGA2共表达基因，在酿酒酵母中构建β-丙氨酸途径。改善前体供应和丙二酸生产。通过对两个异源基因进行基于delta序列的整合，筛选出产7.21 mg/L丙二酸的工程菌株。此外，将琥珀酸半醛脱氢酶基因UGA2替换为来自大肠杆菌的yneI ，该基因在先前的研究中用于生产丙二酸，使烧瓶培养中的丙二酸滴度增加到 7.96 mg/L。经过优化，最终工程菌株在摇瓶中发酵产生的最大丙二酸滴度为 12.83 mg/L，并且在 5 L 生物反应器中的分批补料发酵过程中增加到 91.53 mg/L，与之前相比增加了两倍。过表达UGA2的工程菌株。