枯草芽孢杆菌ZK8产伊枯草菌素A发酵过程中的氮代谢  

Nitrogen metabolism from Bacillus subtilis ZK8 during Iturin A fermentation

作　　者：庞琰 岳华 钟娟[1] 舒丹[1] 罗笛[1] 杨杰[1] 周金燕[1] 李哲敏 谭红[1] PANG Yan;YUE Hua;ZHONG Juan;SHU Dan;LUO Di;YANG Jie;ZHOU Jinyan;LI Zhemin;TAN Hong(CAS Key Laboratory of Environmental and Applied Microbiology,Chengdu Institute of Biology,Chinese Academy of Sciences,Chengdu 610041,China;University of Chinese Academy of Sciences,Beijing 100049,China)

机构地区：[1]中国科学院成都生物研究所,中国科学院环境与应用微生物重点实验室,成都610041 [2]中国科学院大学,北京100049

出　　处：《应用与环境生物学报》2022年第3期553-560,共8页Chinese Journal of Applied and Environmental Biology

基　　金：中国科学院“西部之光”青年博士B类项目(2017XBZG_XBQNXZ_B_006);中国科学院环境与应用微生物重点实验室开放基金(KLCAS-2018-3)资助。

摘　　要：在分批发酵和氨基酸蛋白胨补料分批发酵两种工艺条件下,对枯草芽孢杆菌(Bacillus subtilis)ZK8产伊枯草菌素A(Iturin A)发酵过程中的胞外中性蛋白酶、细胞内外游离氨基酸、蛋白质及总氮含量进行测定及规律分析.结果表明,相较于分批发酵工艺,补料分批发酵工艺条件下胞外中性蛋白酶活力受到抑制,且在发酵中后期含量急剧下降;两种工艺条件下18种胞外游离氨基酸含量变化规律差别较大,分批发酵过程中有15种氨基酸在发酵开始第6或12小时便迅速耗尽,而补料发酵工艺使氨基酸供给相对充足;补料分批发酵过程中胞外蛋白质含量都呈现积累的趋势;至60 h发酵结束,分批发酵和补料分批发酵工艺条件下,Iturin A的氮源转化率分别为3.6%和5.13%.结果发现通过氨基酸蛋白胨补料优化了培养基中的氨基酸组成结构,使更多氮元素流向Iturin A合成方向,使Iturin A产量由0.6 g/L提高至1.64 g/L.本研究从氮代谢水平揭示了氨基酸蛋白胨补料分批发酵工艺下B.subtilis ZK8高产Iturin A的机制,可为后续发酵过程精细化调控及菌种代谢工程改良提供理论依据.(图6参25)This study aimed to reveal the high yield mechanism of Iturin A by Bacillus subtilis ZK8 in fed-batch fermentation of amino acid peptone at the metabolic level.Extracellular neutral protease and intracellular and extracellular levels of free amino acids,proteins,and total nitrogen were determined and analyzed under the two process conditions of batch fermentation and amino acid peptone fed-batch fermentation.The results showed that the extracellular neutral protease activity was inhibited in the fed-batch fermentation process compared with that in the batch fermentation process,and the content decreased sharply in the middle and late fermentation stages.Under the two process conditions,the variations in 18 extracellular free amino acids were quite different.During the batch fermentation process,15 amino acids were rapidly exhausted in the 6th or 12th hour,whereas the fed-batch fermentation process provided relatively sufficient amino acids.During fed-batch fermentation,the extracellular protein content showed an accumulation trend.At the end of 60 h of fermentation,the nitrogen source conversion of Iturin A was 3.6%and 5.13%in batch and fed-batch fermentations,respectively.Amino acid composition and structure in the medium were found to be optimized by amino acid peptone feeding,and more nitrogen elements flowed in the direction of Iturin A synthesis;therefore,the yield of Iturin A was further increased from 0.6 g/L to 1.64 g/L.This study reveals the high yield mechanism of Iturin A by B.subtilis ZK8 in fed-batch fermentation of amino acid peptone at the nitrogen metabolic level,which provides a theoretical basis for the finescale regulation of the subsequent fermentation process and the improvement of strain metabolic engineering.

关 键 词：枯草芽孢杆菌 Iturin A 补料分批发酵 代谢分析 发酵过程 

分 类 号：TQ920.6[轻工技术与工程—发酵工程]