溶氧量对酿酒酵母及其工程菌的β-苯乙醇合成代谢的影响及调控效应  

Effects and Biosynthetic Metabolism Regulation of Dissolved Oxygen on β-Phenethylalcohol in Saccharomyces cerevisiae and Its Engineering Strains

作　　者：王成涛[1,2] 梁婧如 尹胜[1] 周娣 孙宝国[1,2] 赵吉兴 

机构地区：[1]北京工商大学北京食品营养与人类健康高精尖创新中心 [2]北京市食品添加剂工程技术研究中心食品质量与安全北京实验室,北京100048 [3]山东中惠食品有限公司,山东惠民251706

出　　处：《中国食品学报》2016年第8期78-86,共9页Journal of Chinese Institute Of Food Science and Technology

基　　金：国家自然科学基金项目(31571801);北京市自然科学基金项目(5122008);北京市科技计划项目(Z151100001215008);科技创新服务能力建设-科技成果转化-提升计划项目(PXM2016-014213-000034)项目

摘　　要：目的：研究溶氧量对酿酒酵母S288C及工程菌ARO8-10的β-苯乙醇合成代谢的影响及调控效应。方法：在5 L发酵罐中装液量60%,控制p H 5.0,温度30℃,通气比0.5～2.0 vvm,定时取样检测发酵液生物量、β-苯乙醇、L-苯丙氨酸转化率以及菌体相关代谢酶活性等。结果：通气条件在0.5～2.0 vvm范围时,野生型菌株S288C的生物量、异柠檬酸脱氢酶（ICDH）酶活力明显高于转氨酶基因ARO8与脱羧酶基因ARO10耦合过表达工程菌ARO8-10;ARO8-10的β-苯乙醇发酵产量、L-苯丙氨酸转氨酶、苯丙酮酸脱羧酶、G6PD、GK及PDC酶活力均明显高于S288C（P≤0.01）。在1.0 vvm通气条件发酵60 h时,ARO8-10的发酵液中葡萄糖已基本用尽,β-苯乙醇产量达到2.68 g/L,L-苯丙氨酸转化率达到47.3%,较S288C分别提高了44.4%和12.4%。1.0 vvm为β-苯乙醇发酵较适宜的通气条件。结论 ：酿酒酵母的β-苯乙醇代谢与艾利希途径、莽草酸途径、EMP、PPP途径TCA循环等密切相关,构成其合成代谢网络,代谢途经多种酶相互影响;ARO8和ARO10基因耦合过表达,增强了L-苯丙氨酸氨基转移酶和苯丙酮脱羧酶活力,有利于提高G6PD、GK及PDC酶活力,β-苯乙醇代谢流量及产量。Objective： β-phenylethanol is an aromatic alcohol with rose scent, and it is commonly used to make many types of edible flavors. In this work, regulation effect of dissolved oxygen（DO） on the β-phenethylalcohol anabolism were investigated for the wild type strain S. cerevisiae S288 C and its engineered strains ARO8-10. Method： With the conditions of 5 L fermentation tank containing fluid volume by 60%, p H 5.0, temperature 30 ℃, air flow 0.5-2vvm and supplementary 16 g glucose, 7g L-Phe at the point-in-time of 12, 24, 36, 48 h in a 5 L automatic biomasses, β-phenethylalcohol yields, L-Phe conversion rates, the activities of transaminase and decarboxylase were all elevated in S288 C and ARO8-10 in the 60 h fermentation course. Results： The S288 C displayed obviously improved production of the biomass, ICDH activity than ARO8 in the air flow 0.5-2vvm. Compared with S288 C, ARO8-10 showed coupling over-expression of aminotransferase gene ARO8 and decarboxylase gene ARO10, had significantly elevated β-phenethylalcohol yield, activities of aminotransferase, decarboxylase, G6 PD, GK and PDC（P≤0.01）, and ARO8-10 had significantly increased β-phenethylalcohol production and L-phenylalanine conversion at 2.68 g/L and 47.3%, respectively, with the optimum conditions of air flow 1vvm at 60 h fermentation, which were 44.4% and 12.4% higher, respectively, than the wild type, and glucose in ARO8-10 fermentation broth had been exhausted. Conclusion： DO of fermented broth was a key influencing factor of biological fermentation, and 1.0 vvm was suitable ventilation conditions for β-phenethylalcohol fermentation. Ehrlich pathway, Shikimic acid pathway, EMP, PPP, TCA cycle were closely related to β-phenethylalcohol metabolism, which constituted β-phenethylalcohol synthetic metabolic network in S. cerevisiae. The ARO8 and ARO10 genes and their expression products play important regulatory roles by enhancing their expression in S. cerevisiae β-phenethylalcohol anabolism, which as a result facilitates acti

关 键 词：酿酒酵母 β-苯乙醇合成代谢调控 溶氧量 关键酶活力 代谢流量 

分 类 号：TS261.11[轻工技术与工程—发酵工程]