caa a22 4500 605505012 CHVBK 20210128100619.0 cr unu---uuuuu 210128e20150101xx s 000 0 eng 10.1007/s00253-014-6175-8 doi (NATIONALLICENCE)springer-10.1007/s00253-014-6175-8 Novel helper factors influencing recombinant protein production in Pichia pastoris based on proteomic analysis under simulated microgravity [Elektronische Daten] [Jie Huangfu, Feng Qi, Hongwei Liu, Hanfa Zou, Muhammad Ahmed, Chun Li] Microgravity and simulated microgravity (SMG) have quite significant effects on numerous microbial cellular processes. The effects of SMG on the production of recombinant proteins and transcription profiling in prokaryotic and eukaryotic expression host have been investigated. The present study showed that SMG significantly enhanced the specific productivities and activities of the reporter enzymes PGUS and AtXYN that were expressed in recombinant Pichia pastoris. Proteomic profiling revealed that 21 proteins were significantly up-regulated and 35 proteins were drastically down-regulated at the stationary phase, when the recombinant P. pastoris responded to SMG. Six strongly up-regulated genes, TPX, FBA, PGAM, ENO, SBA1, and AKR-E, involved in the oxidative stress response, methanol metabolism, glycolytic pathway, and protein folding, were selected to analyze their impacts on recombinant protein production by co-overexpression in the shaker flask fermentation. The co-overexpressed strains, particularly TPX, FBA, and PGAM, demonstrated promising results with approximately 2.46-fold, 1.58-fold, and 1.33-fold increases in the specific yields of PGUS compared to the control after 48h of methanol induction, respectively. In the meantime, the corresponding PGUS specific activities were increased by 2.33-fold, 2.09-fold, and 1.32-fold, respectively. Thiol peroxidase (TPX), which is involved in the oxidative stress response, significantly influenced the transcriptional levels of the reporter gene PGUS. The present study provides valuable information for further exploration of the molecular mechanism of P. pastoris response to SMG and facilitates simulated microgravity for finding novel helper factors to rationally engineer the strains in normal fermentation by using proteomic studies. Springer-Verlag Berlin Heidelberg, 2014 Recombinant protein nationallicence Strengthen production nationallicence Proteomics analysis nationallicence Simulate microgravity (SMG) nationallicence Pichia pastoris nationallicence Huangfu Jie School of Life Science, Beijing Institute of Technology, 100081, Beijing, China aut Qi Feng School of Life Science, Beijing Institute of Technology, 100081, Beijing, China aut Liu Hongwei National Chromatography R&A Centre, Dalian Institute of Chemical Physics, CAS, 116023, Dalian, China aut Zou Hanfa National Chromatography R&A Centre, Dalian Institute of Chemical Physics, CAS, 116023, Dalian, China aut Ahmed Muhammad School of Life Science, Beijing Institute of Technology, 100081, Beijing, China aut Li Chun School of Life Science, Beijing Institute of Technology, 100081, Beijing, China aut Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/2(2015-01-01), 653-665 0175-7598 99:2<653 2015 99 253 https://doi.org/10.1007/s00253-014-6175-8 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s00253-014-6175-8 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Huangfu Jie School of Life Science, Beijing Institute of Technology, 100081, Beijing, China aut NATIONALLICENCE 700 1- Qi Feng School of Life Science, Beijing Institute of Technology, 100081, Beijing, China aut NATIONALLICENCE 700 1- Liu Hongwei National Chromatography R&A Centre, Dalian Institute of Chemical Physics, CAS, 116023, Dalian, China aut NATIONALLICENCE 700 1- Zou Hanfa National Chromatography R&A Centre, Dalian Institute of Chemical Physics, CAS, 116023, Dalian, China aut NATIONALLICENCE 700 1- Ahmed Muhammad School of Life Science, Beijing Institute of Technology, 100081, Beijing, China aut NATIONALLICENCE 700 1- Li Chun School of Life Science, Beijing Institute of Technology, 100081, Beijing, China aut NATIONALLICENCE 773 0- Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/2(2015-01-01), 653-665 0175-7598 99:2<653 2015 99 253