Chen Hailong, Zhu Nianqing, Wang Yan, Gao Xinxin, Song Yuhe, Zheng Jia, Peng Jiaping, Zhang Xin
Jiangsu Key Laboratory of Chiral Pharmaceuticals Biosynthesis, College of Pharmacy and Chemistry & Chemical Engineering, Taizhou University, 93 Ji Chuan Road, Taizhou, 225300, People's Republic of China.
AMB Express. 2021 Jan 19;11(1):20. doi: 10.1186/s13568-021-01179-8.
Reprogramming glycolysis for directing glycolytic metabolites to a specific metabolic pathway is expected to be useful for increasing microbial production of certain metabolites, such as amino acids, lipids or considerable secondary metabolites. In this report, a strategy of increasing glycolysis by altering the metabolism of inositol pyrophosphates (IPs) for improving the production of S-adenosyl-L-methionine (SAM) for diverse pharmaceutical applications in yeast is presented. The genes associated with the metabolism of IPs, arg82, ipk1 and kcs1, were deleted, respectively, in the yeast strain Saccharomyces cerevisiae CGMCC 2842. It was observed that the deletions of kcs1 and arg82 increased SAM by 83.3 % and 31.8 %, respectively, compared to that of the control. In addition to the improved transcription levels of various glycolytic genes and activities of the relative enzymes, the levels of glycolytic intermediates and ATP were also enhanced. To further confirm the feasibility, the kcs1 was deleted in the high SAM-producing strain Ymls1ΔGAPmK which was deleted malate synthase gene mls1 and co-expressed the Acetyl-CoA synthase gene acs2 and the SAM synthase gene metK1 from Leishmania infantum, to obtain the recombinant strain Ymls1Δkcs1ΔGAPmK. The level of SAM in Ymls1Δkcs1ΔGAPmK reached 2.89 g L in a 250-mL flask and 8.86 g L in a 10-L fermentation tank, increasing 30.2 % and 46.2 %, respectively, compared to those levels in Ymls1ΔGAPmK. The strategy of increasing glycolysis by deletion of kcs1 and arg82 improved SAM production in yeast.
重编程糖酵解以将糖酵解代谢物导向特定代谢途径，有望用于提高某些代谢物的微生物产量，如氨基酸、脂质或大量次生代谢物。在本报告中，提出了一种通过改变肌醇焦磷酸（IPs）代谢来增加糖酵解，以提高酵母中用于多种药物应用的S-腺苷-L-甲硫氨酸（SAM）产量的策略。分别在酿酒酵母CGMCC 2842菌株中删除了与IPs代谢相关的基因arg82、ipk1和kcs1。观察到，与对照相比，kcs1和arg82的缺失分别使SAM增加了83.3%和31.8%。除了各种糖酵解基因的转录水平提高和相关酶的活性增强外，糖酵解中间体和ATP的水平也有所提高。为了进一步证实其可行性，在高SAM产生菌株Ymls1ΔGAPmK（该菌株删除了苹果酸合酶基因mls1并共表达了来自婴儿利什曼原虫的乙酰辅酶A合酶基因acs2和SAM合酶基因metK1）中删除kcs1，以获得重组菌株Ymls1Δkcs1ΔGAPmK。在250 mL摇瓶中，Ymls1Δkcs1ΔGAPmK中的SAM水平达到2.89 g/L，在10 L发酵罐中达到8.86 g/L，与Ymls1ΔGAPmK中的水平相比，分别提高了30.2%和46.2%。通过删除kcs1和arg82来增加糖酵解的策略提高了酵母中SAM的产量。
