Metabolic Engineering of the Regulators in Nitrogen Catabolite Repression To Reduce the Production of Ethyl Carbamate in a Model Rice Wine System
Metabolic Engineering of the Regulators in Nitrogen Catabolite Repression To Reduce the Production of Ethyl Carbamate in a Model Rice Wine System
复制标题
氮分解代谢物抑制调节剂的代谢工程，以减少模型米酒系统中氨基甲酸乙酯的产生
DOI：
10.1128/aem.03055-13
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发表时间：
2014-01-01
影响因子：
4.4
通讯作者：
Chen, Jian
中科院分区：
文献类型：
--
作者：
Zhao, Xinrui;Zou, Huijun;Chen, Jian
黄酒一直是中国最受欢迎的传统酒类之一。然而，黄酒中潜在致癌物质氨基甲酸乙酯(EC)的存在引发了一系列食品安全问题。在黄酒生产过程中，酿酒酵母因氮分解代谢抑制(NCR)而产生的尿素积累是形成EC的关键原因。当存在较好的氮源时，NCR通过将Gln3p保留在细胞质中来抑制尿素的利用。为了增加Gln3p的核定位，核定位信号上的一些可能的磷酸化位点发生了突变，核定位调节信号被截断，URE2的破坏提供了另一种减少尿素积累的方法。通过结合这些策略，参与尿素利用的基因(DUR1，2和DUR3)可以在谷氨酰胺存在的情况下显著激活。在转基因菌株的摇瓶发酵过程中，几乎没有尿素在培养基中积累。此外，在模型黄酒体系中，尿素和EC的浓度分别降低了63%和72%。对黄酒中正常营养成分的检测表明，野生菌株和转基因菌株在发酵特性上差异不大。这些结果表明，NCR调节剂的代谢工程作为一种消除黄酒生产中的EC的方法具有很大的潜力。
Rice wine has been one of the most popular traditional alcoholic drinks in China. However, the presence of potentially carcinogenic ethyl carbamate (EC) in rice wine has raised a series of food safety issues. During rice wine production, the key reason for EC formation is urea accumulation, which occurs because of nitrogen catabolite repression (NCR) in Saccharomyces cerevisiae. NCR represses urea utilization by retaining Gln3p in the cytoplasm when preferred nitrogen sources are present. In order to increase the nuclear localization of Gln3p, some possible phosphorylation sites on the nuclear localization signal were mutated and the nuclear localization regulation signal was truncated, and the disruption of URE2 provided an additional method of reducing urea accumulation. By combining these strategies, the genes involved in urea utilization (DUR1,2 and DUR3) could be significantly activated in the presence of glutamine. During shake flask fermentations of the genetically modified strains, very little urea accumulated in the medium. Furthermore, the concentrations of urea and EC were reduced by 63% and 72%, respectively, in a model rice wine system. Examination of the normal nutrients in rice wine indicated that there were few differences in fermentation characteristics between the wild-type strain and the genetically modified strain. These results show that metabolic engineering of the NCR regulators has great potential as a method for eliminating EC during rice wine production.