Wu Xuefeng, Yao Hongli, Cao Lili, Zheng Zhi, Chen Xiaoju, Zhang Min, Wei Zhaojun, Cheng Jieshun, Jiang Shaotong, Pan Lijun, Li Xingjiang
School of Food Science and Engineering, Hefei University of TechnologyHefei, China.
Key Laboratory for Agricultural Products Processing of Anhui Province, Hefei University of TechnologyHefei, China.
Front Microbiol. 2017 Sep 6;8:1713. doi: 10.3389/fmicb.2017.01713. eCollection 2017.
Pyrroquinoline quinone-dependent alcohol dehydrogenase (PQQ-ADH) is a key enzyme in the ethanol oxidase respiratory chain of acetic acid bacteria (AAB). To investigate the effect of PQQ-ADH on acetic acid production by JST-S, subunits I (adhA) and II (adhB) of PQQ-ADH were over-expressed, the fermentation parameters and the metabolic flux analysis were compared in the engineered strain and the original one. The acetic acid production was improved by the engineered strain (61.42 g L) while the residual ethanol content (4.18 g L) was decreased. Analysis of 2D maps indicated that 19 proteins were differently expressed between the two strains; of these, 17 were identified and analyzed by mass spectrometry and two-dimensional gel electrophoresis. With further investigation of metabolic flux analysis (MFA) of the pathway from ethanol and glucose, the results reveal that over-expression of PQQ-ADH is an effective way to improve the ethanol oxidation respiratory chain pathway and these can offer theoretical references for potential mechanism of metabolic regulation in AAB and researches with its acetic acid resistance.
吡咯喹啉醌依赖型乙醇脱氢酶(PQQ-ADH)是醋酸菌(AAB)乙醇氧化呼吸链中的关键酶。为研究PQQ-ADH对JST-S产乙酸的影响,对PQQ-ADH的亚基I(adhA)和亚基II(adhB)进行了过表达,并比较了工程菌株和原始菌株的发酵参数及代谢通量分析。工程菌株的乙酸产量提高(61.42 g/L),而残余乙醇含量(4.18 g/L)降低。二维图谱分析表明,两菌株之间有19种蛋白质表达存在差异;其中,17种通过质谱和二维凝胶电泳进行了鉴定和分析。通过对乙醇和葡萄糖途径的代谢通量分析(MFA)进一步研究发现,PQQ-ADH的过表达是改善乙醇氧化呼吸链途径的有效方法,可为醋酸菌代谢调控的潜在机制及其耐乙酸研究提供理论参考。