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Comparative transcriptome analysis of global effect of ddh and lysE deletion on 4-hydroxyisoleucine production in corynebacterium glutamicum

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Systems Microbiology and Biomanufacturing 2022年第3期2卷 542-554页

作者： Rui Chen Youhe Xiang Xinping Yu Meiling Zhang Feng Shi State Key Laboratory of Food Science and Technology Jiangnan University1800 Lihu AvenueWuxi 214122China Key Laboratory of Industrial Biotechnology Ministry of EducationSchool of BiotechnologyJiangnan UniversityWuxi 214122China International Joint Laboratory On Food Safety Jiangnan UniversityWuxi 214122China

4-hydroxyisoleucine(4-HIL)is a potential drug for diabetes and weight control.4-HIL was produced by expressing ido gene in L-isoleucine(Ile)-producing corynebacterium *** L-lysine(Lys)was also accumulated as the main by-product in this recombinant strain *** attenuate Lys synthesis,two genes in Lys synthetic pathway,i.e.,ddh encoding the diaminopimelic acid dehydrogenase and lysE encoding the specific Lys exporter were deleted in ***,the deletion of ddh increased 4-HIL titer by 28.1%,but did not decrease Lys content;while the deletion of lysE significantly reduced Lys content by 66.7%,but 4-HIL titer also decreased by 19.3%.Therefore,we carried out transcriptome analysis to reveal the global variation in these *** of ddh and lysE(especially lysE)enhanced the transcription of key enzymes in succinylase branch of Lys synthesis pathway(DapD and DapC)and several enzymes involved in succinyl-CoA accessibility(SucC,SucD and OdhI),suggesting the compensatory synthesis of Lys via succinylase *** addition,the transcription of ilvBN in Ile synthesis pathway was improved,while the transcription of some genes in the 2-methylcitrate cycle and inositol metabolism pathway was weakened in these *** deletion of ddh enhanced the transcription of aceA,ppc and pck,thus promoting oxaloacetate supply and 4-HIL *** of lysE affected the transcription of some stress-related genes and transporter genes,suggesting that this mutant would be under stress,thus attenuating its 4-HIL *** findings will be helpful for systematic microbiology and bio-manufacturing of ***.

关键词： 4-hydroxyisoleucine corynebacterium glutamicum Transcriptome L-lysine

Improve gamma‑aminobutyric acid production in corynebacterium glutamicum by optimizing the metabolic fux

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Systems Microbiology and Biomanufacturing 2022年第2期2卷 305-316页

作者： Chengzhen Yao Yuantao Liu Xiaoqing Hu Xiaoyuan Wang State Key Laboratory of Food Science and Technology Jiangnan UniversityWuxi 214122China International Joint Laboratory on Food Safety Jiangnan UniversityWuxiJiangsuChina Key Laboratory of Industrial Biotechnology Ministry of EducationSchool of BiotechnologyJiangnan UniversityWuxiChina Hulunbeier Northeast Fufeng Biotechnology Co.Ltd. HulunbeierChina

Gamma-aminobutyric acid is an important nonprotein amino acid and has been extensively applied in pharmaceuticals,livestock,food additives,and so *** is important to develop corynebacterium glutamicum strains that can efciently produce gamma-aminobutyric acid from *** this study,production of gamma-aminobutyric acid in *** CGY700 was improved by construction of CO_(2) anaplerotic reaction and overexpression of citrate *** co-expression of ppc encoding phosphoenolpyruvate carboxylase and gltA encoding citrate synthase was constructed and optimized in the chromosome to compensate carbon loss and conquer metabolic *** expression of ppc and gltA were controlled by promoters P_(tac) and P_(tacM),and the optimal mode of P_(tacM-ppc)-P_(tac)-gltA was ***,the genes pknG encoding serine/threonine protein kinase G and ldh encoding l-lactate dehydrogenase were deleted,and glnA2 encoding glutamine synthase was overexpressed in the *** fnal strain CGY-PG-304 constructed in this study could produce 41.17 g/L gamma-aminobutyric acid in shake fask cultivation and 58.33 g/L gamma-aminobutyric acid via FedBatch fermentation with a yield of 0.30 g/g ***-PG-304 was constructed by genome editing;therefore,it is stable and not necessary to add any antibiotics and inducer during fermentation.

关键词： Gamma-aminobutyric acid Metabolic engineering corynebacterium glutamicum Phosphoenolpyruvate carboxylase Citrate synthase

Development of a novel platform for recombinant protein production in corynebacterium glutamicum on ethanol

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Synthetic and Systems Biotechnology 2022年第2期7卷 765-774页

作者： Xinyu Yu Xiuxia Liu Xiong Gao Xunxun Luo Yankun Yang Ye Li Chunli Liu Chong Zhang Zhonghu Bai Key Laboratory of Industrial Biotechnology Ministry of EducationSchool of BiotechnologyJiangnan UniversityWuxiChina National Engineering Research Center for Cereal Fermentation and Food Biomanufacturing Jiangnan UniversityWuxiChina Division of Life Science The Hong Kong University of Science and TechnologyClear Water BayHong KongChina Jiangsu Provincial Research Center for Bioactive Product Processing Technology Jiangnan UniversityWuxiChina MOE Key Laboratory for Industrial Biocatalysis Institute of Biochemical EngineeringDepartment of Chemical EngineeringTsinghua UniversityBeijingChina Center for Synthetic and Systems Biology Tsinghua UniversityBeijingChina

corynebacterium glutamicum represents an emerging recombinant protein expression factory due to its ideal features for protein secretion,but its applicability is harmed by the lack of an autoinduction system with tight regulation and high ***,we propose a new recombinant protein manufacturing platform that leverages ethanol as both a delayed carbon source and an ***,we reanalysed the native inducible promoter PICL from the acetate uptake operon and found that its limited capacity is the result of the inadequate translation initial *** two strategies of bicistronic design and ribozyme-based insulator can ensure the high activity of this ***,through transcriptional engineering that alters transcription factor binding sites(TFBSs)and the first transcribed sequence,the truncated promoter PA256 with a dramatically higher transcription level was *** producing the superfolder green fluorescent protein(sfGFP)under 1%ethanol conditions,PA256 exhibited substantially lower protein accumulation in prophase but an approximately 2.5-fold greater final yield than the strong promoter *** superior expression mode was further validated using two secreted proteins,camelid antibody fragment(VHH)and endoxylanase(XynA).Furthermore,utilizing CRISPRi technology,ethanol utilization blocking strains were created,and PA256 was shown to be impaired in the phosphotransacetylase(PTA)knockdown strains,indicating that ethanol metabolism into the tricarboxylic acid cycle is required for PA256 ***,this platform was applied to produce the“de novo design”protein NEO-2/15,and by introducing the N-propeptide of CspB,NEO-2/15 was effectively secreted with the accumulation 281 mg/L obtained after 24 h of shake-flask *** the best of our knowledge,this is the first report of NEO-2/15 secretory overexpression.

关键词： corynebacterium glutamicum Ethanol Transcriptional engineering Secretory protein overexpression NEO-2/15

Small protein Cgl2215 enhances phenolic tolerance by promoting MytA activity in corynebacterium glutamicum

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Stress Biology 2022年第1期2卷 97-108页

作者： Huawei Gu Xinwei Hao Ruirui Liu Zhenkun Shi Zehua Zhao Fu Chen Wenqiang Wang Yao Wang Xihui Shen State Key Laboratory of Crop Stress Biology for Arid Areas Shaanxi Key Laboratory of Agricultural and Environmental MicrobiologyCollege of Life SciencesNorthwest A&F UniversityYanglingShaanxiChina

corynebacterium glutamicum is a promising chassis microorganism for the bioconversion of lignocellulosic biomass owing to its good tolerance and degradation of the inhibitors generated in lignocellulosic *** the identified proteins encoded by genes within the *** genome,nearly 400 are still functionally *** on previous transcriptome analysis,we found that the hypothetical protein gene cgl2215 was highly upregu-lated in response to phenol,ferulic acid,and vanillin *** cgl2215 deletion mutant was shown to be more sensitive than the parental strain to phenolic compounds as well as other environmental factors such as heat,ethanol,and oxidative ***2215 interacts with *** mycoloyltransferase A(MytA)and enhances its in vitro esterase *** assays of theΔmytA andΔcgl2215ΔmytA mutants in response to phenolic stress estab-lished that the role of Cgl2215 in phenolic tolerance was mediated by ***,transmission electron microscopy(TEM)results showed that cgl2215 and mytA deletion both led to defects in the cell envelope structure of ***,especially in the outer layer(OL)and electron-transparent layer(ETL).Collectively,these results indicate that Cgl2215 can enhance MytA activity and affect the cell envelope structure by directly interacting with MytA,thus playing an important role in resisting phenolic and other environmental stresses.

关键词： corynebacterium glutamicum Tolerance Phenolic compounds MytA Mycoloyltransferase Cell envelope

Two-stage pH Control Mode in Batch Fermentation of a Novel Bioflocculant from corynebacterium glutamicum

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Chemical Research in Chinese Universities 2004年第2期20卷 152-155页

作者： HENing WUXiao-jie DENGXu LUYing-hua LIQing-biao DepartmentofChemicalandBiochemicalEngineering CollegeofChemistryandChemicalEngineeringXiamenUniversityXiamen361005P.R.China

The effect of pH of the fermentation medium on cell growth and the production of a novel bioflocculant(named REA-11) by corynebacterium glutamicum CCTCC M201005 were investigated. The maximum biomass(2.23 g/L) and flocculating activity(142.2 U/mL) were simultaneously obtained at the 14th hour when the pH value of the culture medium was maintained at 7.0 during the whole fermentation process. The production of REA-11 kept on a trend of increase till the later phase of fermentation process, which resulted in the ultimate flocculating activity of the culture broth to enhance to nearly 100 U/mL at pH 6.0. A two-stage pH control mode was adopted in REA-11 production in which the pH value of the culture medium was controlled at 7.0 during the first 14 h, then decreased to 6.0 that was maintained until the end of the fermentation process. With the two-stage pH control mode, the maximum flocculating activity reached 178.8 U/mL which was 30% higher than that obtained under the condition of pH 7.0 and the biomass enhanced about 15%. Compared with the fermentation process without pH control, REA-11 production and cell growth via the two-stage pH control mode increased 80% and 25%, respectively.

关键词： corynebacterium glutamicum pH control Bioflocculant

Engineering corynebacterium glutamicum for Geraniol Production

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Transactions of Tianjin University 2021年第5期27卷 377-384页

作者： Man Li Shuo Xu Wenyu Lu School of Chemical Engineering and Technology Tianjin UniversityTianjin 300072China Key Laboratory of System Bioengineering(Tianjin University) Ministry of EducationTianjin 300072China SynBio Research Platform Collaborative Innovation Center of Chemical Science and Engineering(Tianjin)Tianjin 300072China

Geraniol is a monoterpenoid alcohol with various applications in food,cosmetics,and *** glutamicum is a potential platform for terpenoids production because it harbors the methylerythritol phosphate *** engineer *** to produce geraniol,two different truncated geraniol synthases (GESs) were respectively expressed,and strain LX02 expressing the truncated GESs from Valeriana officinalis (t Vo GES) produced 0.3 mg/L of ***,three geranyl diphosphate synthases (GPPSs) were combinatorially co-expressed with t Vo GES to improve geraniol *** amounts of produced geraniol were all higher than that produced by strain *** LX03 co-expressing ERG20 F96W–N127W (ERG20 WW) and t Vo GES produced the highest amount,5.4 mg/***,the co-overexpression of1-deoxy-D-xylulose-5-phosphate synthase (dxs) and isopentenyl diphosphate isomerase (idi) further increased the production to 12.2 mg/L in strain ***,the production of geraniol was increased to 15.2 mg/L via fermentation *** our knowledge,this is the first report on the engineering of *** to produce geraniol and thus can serve as a reference for other monoterpenoid production studies.

关键词： corynebacterium glutamicum Geraniol Geraniol synthase Overexpression Metabolic engineering

An application of dicistronic vector for protein expression in corynebacterium glutamicum

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An application of dicistronic vector for protein expression ...

Characterization and implications of prokaryotic ribosome-binding sites across species

中国生物化学与分子生物学会2016年全国学术会议

作者： Wei Zhang Zihao Zhao Feng Peng Yankun Yang Zhonghu Bai Xiuxia Liu National Engineering Laboratory for Cereal Fermention Technology Jiangnan University The Key Laboratory of Industrial Biotechnology Ministry of Education Jiangnan University

corynebacterium glutamicum is traditionally used in fermentation processes on amino acids production,and it is also an attractive host for the production of heterologous proteins such as industrial enzymes and pharmaceutical ***,attempts to use *** as hsot for heterologous protein production mainly focused on vector construction or promoter screening have often failed or lead to disappointing ***,few expression vectors also limited the research on

关键词： corynebacterium glutamicum dicistronic vector translation reinitiation secretory production

来源： cnki会议评论

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Systems Microbiology and Biomanufacturing 2022年第4期2卷 676-684页

作者： Yanting Duan Xiaojuan Zhang Weiji Zhai Jinpeng Zhang Jiawei Ren Xiaomei Zhang Guoqiang Xu Jinsong Shi Zhenghong Xu Biotechnology of Ministry of Education School of BiotechnologyJiangnan UniversityWuxiChina National Engineering Research Center for Cereal Fermentation and Food Biomanufacturing Jiangnan UniversityWuxiChina School of Life Science and Health Engineering Jiangnan UniversityWuxi 214122China Jiangsu Engineering Research Center for Bioactive Products Processing Technology Jiangnan University1800 Lihu AvenueWuxi 214122China

The ribosome-binding site(RBS)in the 5′untranslated region is recognized by 16S rRNA to start translation and is an essential element of the gene expression *** have been widely applied in regulating gene expression in various scenarios,including Gram-negative or Gram-positive ***,we first rationally designed and constructed an RBS mutant library containing 66 *** strength of these RBSs in *** and *** was characterized *** RBS strength spanned about 200 and 15 times in the two species,*** strength of RBSs in *** was generally lower than that of in ***.A total of 18 RBSs showed similar strength(within twofold differences)between the species in our study,and the correlation analysis of the strength of RBSs between *** and ***(R^(2)=0.7483)revealed that these RBSs can be used across *** sequence analysis revealed that the RBS region with two Ts stated was beneficial for RBS to function *** RBS characterized here can be used to precisely regulate gene expression in both hosts,and the characteristics of cross-species RBSs provide basic information for RBS rational design.

关键词： RBS library Cross-species RBSs Sequence characteristics Escherichia coli corynebacterium glutamicum

Microbial chassis design and engineering for production of amino acids used in food industry

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Systems Microbiology and Biomanufacturing 2023年第1期3卷 28-48页

作者： Jianli Wang Wenjian Ma Jingwen Zhou Xiaoyuan Wang State Key Laboratory of Food Science and Technology Jiangnan University1800 Lihu AvenueWuxi 214000China International Joint Laboratory on Food Safety Jiangnan UniversityWuxi 214122China Science Center for Future Foods Jiangnan UniversityWuxi 214122China Key Laboratory of Industrial Biotechnology Ministry of EducationSchool of BiotechnologyJiangnan UniversityWuxi 214122China

Rational microbial chassis design and engineering for improving production of amino acids have attracted a considerable attention.l-glutamate,l-lysine,l-threonine and l-tryptophan are the main amino acids demanded in the food *** metabolic engineering and synthetic biology engineering generally are believed as the comprehensive engineering approaches to obtain rationally designed strains and construct high-performance platforms for amino *** strate-gies focus on microbial chassis characterization optimization,precise metabolic engineering such as promoter engineer-ing,modular pathway engineering,transporter engineering,and dynamic switch systems application,and global genome streamline engineering to reduce cell *** this review,we summarized the efficient engineering strategies to optimize corynebacterium glutamicum and Escherichia coli cell factories for improving the production of l-glutamate,l-lysine,l-threonine,and l-tryptophan.

关键词： Microbial chassis Amino acids Food industries Synthetic biology Metabolic engineering Escherichia coli corynebacterium glutamicum l-Glutamate l-Lysine l-Threonine l-Tryptophan