文献类型：期刊文献

中文题名：Combined ^(13)C-assisted metabolomics and metabolic flux analysis reveals the impacts of glutamate on the central metabolism of highβ-galactosidase-producing Pichia pastoris

作者：Ping Liu[1];Mingzhi Huang[1];Menglei Guo[1];Jiangchao Qian[1];Weilu Lin[1];Ju Chu[1];Yingping Zhuang[1];Siliang Zhang[1]

机构：[1]State Key Laboratory of Bioreactor Engineering,East China University of Science and Technology,No.130,Meilong Road,Shanghai 200237,China

年份：2016

卷号：3

期号：1

起止页码：67

中文期刊名：Bioresources and Bioprocessing

外文期刊名：生物资源与生物加工(英文)

基金：supported by the National Basic Research Program of China(973 Program)(No.2013CB733600);National Natural Science Foundation of China(No.201276081);National Key Technology Support Program of China(No.2011BAF02B05).

语种：英文

中文关键词：Glutamate;^(13)C metabolic flux analysis;Metabolomics;P.pastoris;Recombinant protein expression

摘要：Background:Pichia pastoris is a popular recombinant protein expression system for its accessibility of efficient gene manipulation and high protein production.Sufficient supply of precursors,energy,and redox cofactors is crucial for high recombinant protein production.In our present work,we found that the addition of glutamate improved the recombinantβ-galactosidase(β-gal)production by P.pastoris G1HL.Methods:To elucidate the impacts of glutamate on the central metabolism in detail,a combined ^(13)C-assisted metabolomics and ^(13)C metabolic flux analysis was conducted based on LC-MS/MS and GC-MS data.Results:The pool sizes of intracellular amino acids were obviously higher on glucose/glutamate(Glc/Glu).The fluxes in EMP entry reaction and in downstream TCA cycle were 50 and 67%higher on Glc/Glu than on Glc,respectively.While the fluxes in upstream TCA cycle kept almost unaltered,the fluxes in PPP oxidative branch decreased.Conclusion:The addition of glutamate leads to a remarkable change on the central metabolism of highβ-galactosidase-producing P.pastoris G1HL.To meet the increased demands of redox cofactors and energy for higherβ-galactosidase production on Glc/Glu,P.pastoris G1HL redistributes the fluxes in central metabolism through the inhibitions and/or activation of the enzymes in key nodes together with the energy and redox status.