生物过程代谢组学与代谢流测定
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摘要
整合的代谢组学和13C代谢流数据能充分反映细胞的代谢状态,对代谢组学和代谢流的测定技术已成为工业生物过程研究的重要手段,并能为工业生物过程优化和高产菌株理性设计提供重要帮助。阐述了代谢组学和代谢流测定的完整流程,包括实验方法、数据处理方法和软件工具,并综述了其在代谢途径鉴定、代谢机制解析和代谢工程等领域的应用进展。
The integration of metabolomics and 13 C metabolic fluxes provide an adequate description of cell metabolic state. The determination of metabolomics and metabolic fluxes is crucial and helpful for the optimization of industrial bioprocesses and rational design of high production strains. This paper illustrates the entire workflows for determining metabolomics and metabolic fluxes, including experiment methods, data processing methods and software tools. The application of metabolomics and metabolic fluxes in the fi elds of metabolic pathway identifi cation, metabolic mechanism elucidation and metabolic engineering is also summarised.
The integration of metabolomics and 13 C metabolic fluxes provide an adequate description of cell metabolic state. The determination of metabolomics and metabolic fluxes is crucial and helpful for the optimization of industrial bioprocesses and rational design of high production strains. This paper illustrates the entire workflows for determining metabolomics and metabolic fluxes, including experiment methods, data processing methods and software tools. The application of metabolomics and metabolic fluxes in the fi elds of metabolic pathway identifi cation, metabolic mechanism elucidation and metabolic engineering is also summarised.
引文
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[14]JING L,LEI Z T,ZHANG G W,et al.Metabolite profiles of essential oils in citrus peels and their taxonomic implications[J].Metabolomics,2015,11(4):952-963.
[15]POLAKOF S,REMOND D,RAMBEAU M,et al.Postprandial metabolic events in mini-pigs:newinsights from a combined approach using plasma metabolomics,tissue gene expression,and enzyme activity[J].Metabolomics,2015,11(4):964-979.
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[23]AHN W S,ANTONIEWICZ M R.Metabolic flux analysis of CHO cells at growth and non-growth phases using isotopic tracers and mass spectrometry[J].Metab Eng,2011,13(5):598-609.
[24]TEMPLETON N,DEAN J,REDDY P,et al.Peak antibody production is associated with increased oxidative metabolism in an industrially relevant fed‐batch CHO cell culture[J].Biotechnol Bioeng,2013,110(7):2013-2024.
[25]LU H,LIU X,HUANG M,et al.Integrated isotope-assisted metabolomics and 13C metabolic flux analysis reveals metabolic flux redistribution for high glucoamylase production by Aspergillus niger[J].Microbial Cell Factories,2015,14(1):1-14.
[26]HU J,LEI P,MOHSIN A,et al.Mixomics analysis of Bacillus subtilis:effect of oxygen availability on ribof lavin production[J].Microbial Cell Factories,2017,16(1):150-165.
[27]AURICH M K,PAGLIA G,ROLFSSON O,et al.Prediction of intracellular metabolic states from extracellular metabolomic data[J].Metabolomics,2015,11(3):603-619.
[28]LU H,CAO W,OUYANG L,et al.Comprehensive reconstruction and in silico analysis of Aspergillus niger genome‐scale metabolic network model that accounts for 1210 ORFs[J].Biotechnol Bioeng,2017,114(3):685-695.
[29]YE R,HUANG M,LU H,et al.Comprehensive reconstruction and evaluation of Pichia pastoris genome-scale metabolic model that accounts for 1243 ORFs[J].Bioresour Bioprocess,2017,4(1):22-33.
[2]VAN GULIK W M.Fast sampling for quantitative microbial metabolomics[J].Curr Opin Biotech,2010,21(1):27-34.
[3]CHENG K K,LEE B S,MASUDA T,et al.Global metabolic network reorganization by adaptive mutations allows fast growth of Escherichia coli on glycerol[J].Nature Communications,2014,5:3233-3241.
[4]WARTH B,PARICH A,BUESCHI C,et al.GC-MS based targeted metabolic prof iling identifies changes in the wheat metabolome following deoxynivalenol treatment[J].Metabolomics,2015,11(3):722-738.
[5]JUNG J Y,OH M K.Isotope labeling pattern study of central carbon metabolites using GC/MS[J].J Chromatogr B,2015,974:101-108.
[6]SONG Y L,ZHANG N,SHI S P,et al.Large-scale qualitative and quantitative character ization of components in Shenf u injection by integrating hydrophilic interaction ch romatography,reversed phase liquid chromatography,and tandem mass spectrometry[J].J Chromatogr A,2015,1407:106-118.
[7]JACOB C C,DERVILLY-PINEL G,BIANCOTTO G,et al.Global urine fi ngerprinting by LC-ESI(+)-HRMS for better characterization of metabolic pathway disruption upon anabolic practices in bovine[J].Metabolomics,2015,11(1):184-197.
[8]RARO M,IBANEZ M,GIL R,et al.Untargeted metabolomics in doping control:detection of new markers of testosterone misuse by ultrahigh performance liquid chromatography coupled to highresolution mass spectrometry[J].Anal Chem,2015,87(16):8373-8380.
[9]SOLLIEC M,ROY L A,SAUVE S.Quantitative performance of liquid chromatography coupled to Q-Exactive high resolution mass spectrometry(HRMS)for the analysis of tetracyclines in a complex matrix[J].Anal Chim Acta,2015,853:415-424.
[10]GODZIEN J,GARCIA M D,MARTINEZ A P,et al.Effect of a nutraceutical treatment on diabetic rats with targeted and CE-MS non-targeted approaches[J].Metabolomics,2013,9(1):S188-S202.
[11]XIA M L,HUANG D,LI S S,et al.Enhanced FK506 production in streptomyces tsukubaensis by rational feeding strategies based on comparative metabolic profi ling analysis[J].Biotechnol Bioeng,2013,110(10):2717-2730.
[12]ZAKHARTSEV M,VIELHAUER O,HORN T,et al.Fast sampling for quantitative microbial metabolomics:new aspects on cold methanol quenching:metabolite co-precipitation[J].Metabolomics,2015,11(2):286-301.
[13]MISRA B B,VAN DER HOOFT J.Updates in metabolomics tools and resources:2014–2015[J].Electrophoresis,2016,37(1):86-110.
[14]JING L,LEI Z T,ZHANG G W,et al.Metabolite profiles of essential oils in citrus peels and their taxonomic implications[J].Metabolomics,2015,11(4):952-963.
[15]POLAKOF S,REMOND D,RAMBEAU M,et al.Postprandial metabolic events in mini-pigs:newinsights from a combined approach using plasma metabolomics,tissue gene expression,and enzyme activity[J].Metabolomics,2015,11(4):964-979.
[16]ZAMBONI N,FENDT S M,RUHL M,et al.13C-based metabolic flux analysis[J].Nat Protocols,2009,4(6):878-892.
[17]ANTONIEWICZ M R.13C metabolic flux analysis:optimal design of isotopic labeling experiments[J].Cur r Opin Biotechnol,2013,24(6):1116-1121.
[18]DROSTE P,MIEBACH S,NIEDENFUHR S,et al.Visualizing multi-omics data in metabolic networks with the software Omix:a case study[J].Biosystems,2011,105(2):154-161.
[19]SWARUP A,LU J,DEWOODY K C,et al.Metabolic network reconstr uction,growth characterization and 13C-metabolic flux analysis of the extremophile Thermus thermophilus HB8[J].Metab Eng,2014,24:173-180.
[20]KLINGNER A,BARTSCH A,DOGS M,et al.Large-scale 13C flux profi ling reveals conservation of the entner-doudoroff pathway as a glycolytic strategy among marine bacteria that use glucose[J].Appl Environ Microbiol,2015,81(7):2408-2422.
[21]NIE Y,HUANG M,LU J,et al.Impacts of highβ-galactosidase expression on central metabolism of recombinant Pichia pastoris GS115 using glucose as sole carbon source via 13C metabolic flux analysis[J].J Biotechnol,2014,187:124-134.
[22]LIU P,HUANG M,GUO M,et al.Combined 13C-assisted metabolomics and metabolic flux analysis reveals the impacts of glutamate on the central metabolism of highβ-galactosidaseproducing Pichia pastoris[J].Bioresources and Bioprocessing,2016,3(1):47-58.
[23]AHN W S,ANTONIEWICZ M R.Metabolic flux analysis of CHO cells at growth and non-growth phases using isotopic tracers and mass spectrometry[J].Metab Eng,2011,13(5):598-609.
[24]TEMPLETON N,DEAN J,REDDY P,et al.Peak antibody production is associated with increased oxidative metabolism in an industrially relevant fed‐batch CHO cell culture[J].Biotechnol Bioeng,2013,110(7):2013-2024.
[25]LU H,LIU X,HUANG M,et al.Integrated isotope-assisted metabolomics and 13C metabolic flux analysis reveals metabolic flux redistribution for high glucoamylase production by Aspergillus niger[J].Microbial Cell Factories,2015,14(1):1-14.
[26]HU J,LEI P,MOHSIN A,et al.Mixomics analysis of Bacillus subtilis:effect of oxygen availability on ribof lavin production[J].Microbial Cell Factories,2017,16(1):150-165.
[27]AURICH M K,PAGLIA G,ROLFSSON O,et al.Prediction of intracellular metabolic states from extracellular metabolomic data[J].Metabolomics,2015,11(3):603-619.
[28]LU H,CAO W,OUYANG L,et al.Comprehensive reconstruction and in silico analysis of Aspergillus niger genome‐scale metabolic network model that accounts for 1210 ORFs[J].Biotechnol Bioeng,2017,114(3):685-695.
[29]YE R,HUANG M,LU H,et al.Comprehensive reconstruction and evaluation of Pichia pastoris genome-scale metabolic model that accounts for 1243 ORFs[J].Bioresour Bioprocess,2017,4(1):22-33.