文献类型：期刊文献

英文题名：Improving cytosolic aspartate biosynthesis increases glucoamylase production in Aspergillus niger under oxygen limitation

作者：Cao, Weiqiang[1];Wang, Guan[1];Lu, Hongzhong[1];Ouyang, Liming[1];Chu, Ju[1];Sui, Yufei[1];Zhuang, Yingping[1]

机构：[1]East China Univ Sci & Technol, State Key Lab Bioreactor Engn, Shanghai, Peoples R China

年份：2020

卷号：19

期号：1

外文期刊名：MICROBIAL CELL FACTORIES

收录：;WOS:【SCI-EXPANDED(收录号:WOS:000523738200001)】；

基金：This work was financially supported by the National Key Research and Development Program of China (2019YFA0904800), the Science and Technology Commission of Shanghai Municipality (Grant no. 19ZR1413600) and the Fundamental Research Funds for the Central Universities (WF1814032, 22221818014).

语种：英文

外文关键词：Aspergillus niger; Glucoamylase; Limited amino acids; Gene overexpression; Multi-omics study

摘要：Background Glucoamylase is one of the most industrially applied enzymes, produced by Aspergillus species, like Aspergillus niger. Compared to the traditional ways of process optimization, the metabolic engineering strategies to improve glucoamylase production are relatively scarce. Results In the previous study combined multi-omics integrative analysis and amino acid supplementation experiment, we predicted four amino acids (alanine, glutamate, glycine and aspartate) as the limited precursors for glucoamylase production in A. niger. To further verify this, five mutants namely OE-ala, OE-glu, OE-gly, OE-asp1 and OE-asp2, derived from the parental strain A. niger CBS 513.88, were constructed respectively for the overexpression of five genes responsible for the biosynthesis of the four kinds of amino acids (An11g02620, An04g00990, An05g00410, An04g06380 and An16g05570). Real-time quantitative PCR revealed that all these genes were successfully overexpressed at the mRNA level while the five mutants exhibited different performance in glucoamylase production in shake flask cultivation. Notably, the results demonstrated that mutant OE-asp2 which was constructed for reinforcing cytosolic aspartate synthetic pathway, exhibited significantly increased glucoamylase activity by 23.5% and 60.3% compared to CBS 513.88 in the cultivation of shake flask and the 5 L fermentor, respectively. Compared to A. niger CBS 513.88, mutant OE-asp2 has a higher intracellular amino acid pool, in particular, alanine, leucine, glycine and glutamine, while the pool of glutamate was decreased. Conclusion Our study combines the target prediction from multi-omics analysis with the experimental validation and proves the possibility of increasing glucoamylase production by enhancing limited amino acid biosynthesis. In short, this systematically conducted study will surely deepen the understanding of resources allocation in cell factory and provide new strategies for the rational design of enzyme production strains.