Contextualized genome-scale model unveils high-order metabolic effects of the specific growth rate and oxygenation level in recombinant Pichia pastoris
Torres, Paulina (Pontificia Universidad Católica de Chile. Departamento de Ingeniería Química y Bioprocesos)
Saa, Pedro A. (Pontificia Universidad Católica de Chile. Departamento de Ingeniería Química y Bioprocesos)
Albiol i Sala, Joan (Universitat Autònoma de Barcelona. Departament d'Enginyeria Química, Biològica i Ambiental)
Ferrer, Pau (Universitat Autònoma de Barcelona. Departament d'Enginyeria Química, Biològica i Ambiental)
Agosin, Eduardo (Pontificia Universidad Católica de Chile. Departamento de Ingeniería Química y Bioprocesos)

Data:	2019
Resum:	Pichia pastoris is recognized as a biotechnological workhorse for recombinant protein expression. The metabolic performance of this microorganism depends on genetic makeup and culture conditions, amongst which the specific growth rate and oxygenation level are critical. Despite their importance, only their individual effects have been assessed so far, and thus their combined effects and metabolic consequences still remain to be elucidated. In this work, we present a comprehensive framework for revealing high-order (i. e. , individual and combined) metabolic effects of the above parameters in glucose-limited continuous cultures of P. pastoris, using thaumatin production as a case study. Specifically, we employed a rational experimental design to calculate statistically significant metabolic effects from multiple chemostat data, which were later contextualized using a refined and highly predictive genome-scale metabolic model of this yeast under the simulated conditions. Our results revealed a negative effect of the oxygenation on the specific product formation rate (thaumatin), and a positive effect on the biomass yield. Notably, we identified a novel positive combined effect of both the specific growth rate and oxygenation level on the specific product formation rate. Finally, model predictions indicated an opposite relationship between the oxygenation level and the growth-associated maintenance energy (GAME) requirement, suggesting a linear GAME decrease of 0. 56 mmol ATP/g per each 1% increase in oxygenation level, which translated into a 44% higher metabolic cost under low oxygenation compared to high oxygenation. Overall, this work provides a systematic framework for mapping high-order metabolic effects of different culture parameters on the performance of a microbial cell factory. Particularly in this case, it provided valuable insights about optimal operational conditions for protein production in P. pastoris.
Ajuts:	Ministerio de Economía y Competitividad CTQ2016-74959-R Agència de Gestió d'Ajuts Universitaris i de Recerca 2014/SGR-452
Drets:	Aquest document està subjecte a una llicència d'ús Creative Commons. Es permet la reproducció total o parcial, la distribució, i la comunicació pública de l'obra, sempre que no sigui amb finalitats comercials, i sempre que es reconegui l'autoria de l'obra original. No es permet la creació d'obres derivades.
Llengua:	Anglès
Document:	Article ; recerca ; Versió publicada
Matèria:	Pichia pastoris ; Experimental design ; Metabolic modelling ; Dissolved oxygen ; Recombinant protein ; Thaumatin
Publicat a:	Metabolic Engineering Communications, Vol. 9 (December 2019) , art. e00103, ISSN 2214-0301

DOI: 10.1016/j.mec.2019.e00103
PMID: 31720218

10 p, 1.9 MB

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