文献类型：期刊文献

英文题名：Impacts of high β-galactosidase expression on central metabolism of recombinant Pichia pastoris GS115 using glucose as sole carbon source via 13C metabolic flux analysis

作者：Nie, Yongsheng[1];Huang, Mingzhi[1];Lu, Junjie[1];Qian, Jiangchao[1];Lin, Weilu[1];Chu, Ju[1];Zhuang, Yingping[1];Zhang, Siliang[1]

机构：[1]E China Univ Sci & Technol, Key Lab Bioreactor Engn, Shanghai 200237, Peoples R China

年份：2014

卷号：187

起止页码：124

外文期刊名：JOURNAL OF BIOTECHNOLOGY

收录：;EI(收录号：20143518109280);WOS:【SCI-EXPANDED(收录号:WOS:000343837000017)】；

基金：This work was financially supported by the National Basic Research Program of China (973 Program) (No. 2013CB733600), National Natural Science Foundation of China (No. 21276081), National Key Technology Support Program of China (No. 2011BAF02B05), and Research Fund for the Doctoral Program of Higher Education of China (No. 20110074110015).

语种：英文

外文关键词：P. pastoris; C-13 metabolic flux analysis; Central metabolism; Heterologous protein expression

摘要：The yeast Pichia pastoris GS115 is a widely used microbial cell factory for the production of heterologous protein. In order to reveal the impacts of high heterologous protein expression on the central metabolism of Pichia pastoris GS115 using glucose as sole carbon source, we engineered a high beta-galactosidase expression strain P. pastoris G1HL and a low expression control strain P. pastoris GHL through controlling the initiation strength of constitutive promoter p GAP. The carbon flux distributions in these two strains were quantified via C-13 metabolic flux analysis. Compared to the control strain, G1HL showed a lower growth rate, a higher flux through glycolysis pathway, a higher flux through pentose phosphate pathway, and a lower flux through by-products secretion pathway. The metabolic flux redistribution in G1HL was thought to compensate the increased redox cofactors and energy demands caused by the high protein expression. Although the fluxes through Krebs cycle in two engineered strains were almost the same, they were significantly lower than those in wild strain. The enhanced expression of p-galactosidase by glutamate supplementation demonstrated the potential of P. pastoris GS115 to catabolize more carbon through the Krebs cycle for even higher protein expression. In conclusion, our work indicates that P. pastoris GS115 can readjusts the central metabolism for higher heterologous protein expression and provides strategies for strain development or process optimization for enhancing production of heterologous protein. (C) 2014 Elsevier B.V. All rights reserved.