Systematic manipulation of glutathione metabolism in Escherichia coli for improved glutathione production.
Systematic manipulation of glutathione metabolism in Escherichia coli for improved glutathione production.
复制标题
系统地操纵大肠杆菌中的谷胱甘肽代谢以提高谷胱甘肽的产量。
DOI：
10.1186/s12934-016-0439-1
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发表时间：
2016-02-16
影响因子：
6.4
通讯作者：
Ye Q
中科院分区：
文献类型：
--
作者：
Zhang J;Quan C;Wang C;Wu H;Li Z;Ye Q
谷胱甘肽（GSH）是一种具有重要生物学性质的非蛋白巯基化合物，广泛应用于医药、食品、化妆品和保健品中。细胞内GSH的含量取决于GSH合成酶的活性和特性、能量和前体物质的供应以及GSH的降解。在这项研究中，基因编码的前体氨基酸降解和糖原形成以及GSH降解相关的酶进行了系统的操作，在大肠杆菌菌株过表达gshF从琥珀酸放线杆菌。操作包括破坏前体降解途径（tnaA和sdaA），消除l-谷胱甘肽降解（ggt和pepT），以及操纵细胞内ATP水平（破坏glgB）。双向电泳结果表明，突变体GshF的表达量明显降低，其中枢代谢和氨基酸代谢发生了明显的变化。进行五突变体ZJ 12345的补料分批培养，其中GshF表达水平增强，并且GSH产量（19.10 mM）和基于添加的L-半胱氨酸的产量（0.76 mmol/mmol）均显著增加。通过阻断L-半胱氨酸、丝氨酸和GSH的降解途径，阻断糖原的形成，GSH的生产效率显著提高。
l-glutathione (GSH) is a non-protein thiol compound with important biological properties and is widely used in pharmaceutical, food, cosmetic and health products. The cellular GSH is determined by the activity and characteristic of GSH-synthesizing enzymes, energy and precursor supply, and degradation of formed GSH. In this study, genes encoding enzymes related to the precursor amino acid degradation and glycogen formation as well as GSH degradation were systematically manipulated in Escherichia coli strains over-expressing gshF from Actinobacillus succinogenes. The manipulation included disrupting the precursor degradation pathways (tnaA and sdaA), eliminating l-glutathione degradation (ggt and pepT), and manipulating the intracellular ATP level (disruption of glgB). However the constructed mutants showed lower levels of GshF expression. 2-D electrophoresis was performed to elucidate the reasons for this discrepancy, and the results indicated obvious changes in central metabolism and amino acid metabolism in the penta-mutant. Fed-batch culture of the penta-mutant ZJ12345 was performed where the GshF expression level was enhanced, and both the GSH production (19.10 mM) and the yield based on added l-cysteine (0.76 mmol/mmol) were significantly increased. By interrupting the degradation pathways of l-cysteine, serine and GSH and blocking glycogen formation, the GSH production efficiency was significantly improved.