[en] In Komagataella phaffii (Pichia pastoris), formate is a recognized alternative inducer to methanol for expression systems based on the AOX1 promoter (pAOX1). By disrupting the formate dehydrogenase encoding FDH1 gene, we converted such a system into a self-induced one, as adding any inducer in the culture medium is no longer requested for pAOX1 induction. In cells, formate is generated from serine through the THF-C1 metabolism, and it cannot be converted into carbon dioxide in a FdhKO strain. Under non-repressive culture conditions, such as on sorbitol, the intracellular formate generated from the THF-C1 metabolism is sufficient to induce pAOX1 and initiate protein synthesis. This was evidenced for two model proteins, namely intracellular eGFP and secreted CalB lipase from C. antarctica. Similar protein productivities were obtained for a FdhKO strain on sorbitol and a non-disrupted strain on sorbitol-methanol. Considering a K. Phaffii FdhKO strain as a workhorse for recombinant protein synthesis paves the way for the further development of methanol-free processes in K. phaffii.
Disciplines :
Biotechnology
Author, co-author :
Bustos Cosios, Cristina Veronica ; Université de Liège - ULiège > TERRA Research Centre ; School of Biochemical Engineering, Pontificia Universidad Católica de Valparaíso, Valparaiso, Chile
Berrios, Julio ; School of Biochemical Engineering, Pontificia Universidad Católica de Valparaíso, Valparaiso, Chile
Fickers, Patrick ; Université de Liège - ULiège > TERRA Research Centre > Microbial technologies
Language :
English
Title :
Formate from THF-C1 metabolism induces the AOX1 promoter in formate dehydrogenase-deficient Komagataella phaffii.
Publication date :
October 2024
Journal title :
Microbial Biotechnology
ISSN :
1751-7915
eISSN :
1751-7907
Publisher :
John Wiley and Sons Ltd, United States
Volume :
17
Issue :
10
Pages :
e70022
Peer reviewed :
Peer Reviewed verified by ORBi
Funders :
WBI - Wallonie-Bruxelles International Universidad Técnica Federico Santa María FONDECYT - Chile Fondo Nacional de Desarrollo Científico y Tecnológico PUCV - Pontificia Universidad Católica de Valparaíso ANID - Agencia Nacional de Investigación y Desarrollo
Funding text :
The authors thank Prof. Gasser from CD Laboratory of growth-decoupled protein production in yeast, Department of Biotechnology, University of Natural Resources and Life Sciences for providing pKTAC-CRE vector. A. Anckaert, M. Delvenne, Vandenbroucke, V., S. Steels and R. Thomas are acknowledged for their technical help and fruitful discussion. This research was funded by Becas Doctorado Nacional grant number 21211138-Agencia Nacional de Investigaci\u00F3n y Desarrollo (ANID), Chile; Doctoral Internship Scholarship (PUCV, Chile); Research Stay Scholarship N\u00B0 018/2022 (Direcci\u00F3n de Postgrado y Programas, UTFSM, Chile); Wallonie-Bruxelles International through the Cooperation bilateral Belgique-Chili project SUB/2019/435787 (RIO4) and SUB/2023/591923/MOD (RI06), FONDECYT Regular (project number 1191196), University of Liege, Terra Teaching and Research Center.
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