基于galP过表达解除底盘微生物碳抑制效应以提升2,3-BD发酵产率  

Alleviating carbon catabolite repression in the chassis microorganism to enhance fermentation yield of 2,3⁃butanediol based on galP overexpression

作　　者：唐璇 毛亮阳[1,2,3,4] 李娜 杨彤 葛菁萍[1,2,3,4] TANG Xuan;MAO Liangyang;LI Na;YANG Tong;GE Jingping(Heilongjiang University,College of Life Sciences,Harbin 150080,China;Heilongjiang University,Engineering Research Conter of Agricultural Microbial Technology,Ministry of Education,Harbin 150080,China;Heilongjiang University,Heilongjiang Provincial Key Laboratory of Plant Genetic Engineering and Biological Fergineering for Cold Regions,Harbin 150080,China;Heilongjiang University,Key Laboratory of Microbiology,Collgeg of Heilongjiang Province,Harbin 150080,China)

机构地区：[1]黑龙江大学生命科学学院,哈尔滨150080 [2]黑龙江大学农业微生物技术教育部工程研究中心,哈尔滨150080 [3]黑龙江大学黑龙江省寒区植物基因与生物发酵重点实验室,哈尔滨150080 [4]黑龙江大学黑龙江省普通高校微生物重点实验室,哈尔滨150080

出　　处：《黑龙江大学工程学报(中英俄文)》2024年第4期96-105,2,共11页Journal of Engineering of Heilongjiang University

基　　金：国家自然科学基金项目(32071519);黑龙江省生态环境厅2022年度生态环境保护科研项目(HST2022TR004);黑龙江省高校基本科研业务费专项资助项目(2023-KYYWF-1448);功能无机材料化学教育部重点实验室开放课题

摘　　要：为提高肺炎克雷伯氏菌(Klebsiella pneumoniae)有效利用混合碳源的能力及完善ptsG基因缺失对葡萄糖利用的不足,通过基因工程手段,拟进行过表达半乳糖透性酶基因(galP)的方式,形成可高产2,3-丁二醇(2,3-butanediol,简称2,3-BD)的菌株。结果表明,galP得到成功克隆并过表达在K.pneumoniae HD79-N(ΔptsG)菌株后,获得具有既减弱了碳分解代谢抑制(Carbon catabolite repression, CCR)效应,又能够同步利用葡萄糖与木糖生产2,3-BD且终产量达13.15±0.66 g·L^(-1)的工程菌株K.pneumoniae HD79-A。该菌株相较于底盘微生物,2,3-BD终产量提高了34.05%,葡萄糖利用率(1.104 g·L^(-1)·h^(-1))提高了105.59%,总糖消耗量(70.76 g·L^(-1))增加了9.60%。同时,K.pneumoniae HD79-A菌株的生产强度、糖醇转化率以及单位细胞的转化效率分别是K.pneumoniae HD79-N菌株的1.34倍、1.22倍和1.04倍。试验证实了过表达galP能够对菌株K.pneumoniae HD79-N(ΔptsG)进行双重调节,为在混合碳源情况下利用功能性工程菌株高效生产2,3-BD提供理论依据。To enhance capability of Klebsiella pneumoniae in utilizing mixed carbon sources and to address the deficiencies in glucose utilization due to ptsG gene deletion,the genetic engineering to overexpress galactose permease gene(galP)was employed this study.As a result,a strain capable of high 2,3⁃butanediol(2,3⁃BD)production was developed.Successful cloning and overexpression of galP in the K.pneumoniae HD79⁃N(ΔptsG)strain led to the engineered strain K.pneumoniae HD79⁃A.It exhibited reduced carbon catabolite repression(CCR)and the ability to simultaneously utilize glucose and xylose,achieving a final yield of 13.15±0.66 g·L^(-1) of 2,3⁃BD.Compared to the chassis microorganism,the yield of 2,3⁃BD increased by 34.05%,glucose utilization rate(1.104 g·L^(-1)·h^(-1))improved by 105.59%,and total sugar consumption(70.76 g·L^(-1))rose by 9.60%.Additionally,K.pneumoniae HD79⁃A demonstrated that production intensity,alcohol conversion rate,and cell⁃specific conversion efficiency were 1.34⁃fold,1.22⁃fold,and 1.04⁃fold greater than those of K.pneumoniae HD79⁃N,respectively.This study confirmed that overexpressing the galP can effectively dual⁃regulate K.pneumoniae HD79⁃N(ΔptsG),providing a theoretical basis for the efficient production of 2,3⁃BD using functional engineered strains under mixed carbon source conditions.

关 键 词：2 3-丁二醇 过表达 酶基因(galP) 肺炎克雷伯氏菌 

分 类 号：TQ923[轻工技术与工程—发酵工程]