Glycerol metabolism of Pichia pastoris (Komagataella spp.) characterised by 13 C-based metabolic flux analysis
Tomàs Gamisans, Màrius (Universitat Autònoma de Barcelona. Departament d'Enginyeria Química, Biològica i Ambiental)
Ødum, Anders Sebastian Rosenkrans (Technical University of Denmark. Department of Biotechnology and Biomedicine)
Workman, Mhairi (Novo Nordisk)
Ferrer, Pau (Universitat Autònoma de Barcelona. Departament d'Enginyeria Química, Biològica i Ambiental)
Albiol i Sala, Joan (Universitat Autònoma de Barcelona. Departament d'Enginyeria Química, Biològica i Ambiental)

Date:	2019
Abstract:	Metabolic flux analysis based on 13 C-derived constraints has proved to be a powerful method for quantitative physiological characterisation of one of the most extensively used microbial cell factory platforms, Pichia pastoris (syn. Komagataella spp. ). Nonetheless, the reduced number of carbon atoms and the symmetry of the glycerol molecule has hampered the comprehensive determination of metabolic fluxes when used as the labelled C-source. Moreover, metabolic models typically used for 13 C-based flux balance analysis may be incomplete or misrepresent the actual metabolic network. To circumvent these limitations, we reduced the genome-scale metabolic model iMT1026-v3. 0 into a core model and used it for the iterative fitting of metabolic fluxes to the measured mass isotope distribution of proteinogenic amino acids obtained after fractional 13 C labelling of cells with [1,3- 13 C]-glycerol. This workflow allows reliable estimates to be obtained for in vivo fluxes in P. pastoris cells growing on glycerol as sole carbon source, as well as revising previous assumptions concerning its metabolic operation, such as alternative metabolic branches, calculation of energetic parameters and proposed specific cofactor utilisation.
Grants:	Ministerio de Economía y Competitividad CTQ2013-42391-R Ministerio de Economía y Competitividad CTQ2016-74959-R Agència de Gestió d'Ajuts Universitaris i de Recerca 2014/SGR-452 Ministerio de Educación, Cultura y Deporte FPU12/06185
Rights:	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.
Language:	Anglès
Document:	Article ; recerca ; Versió acceptada per publicar
Subject:	C-13-based meabolic flux analysis ; Pichia pastoris ; Komagataella spp ; Glycerol ; Genome-scale metabolic model
Published in:	New Biotechnology, Vol. 50 (May 2019) , p. 52-59, ISSN 1876-4347

DOI: 10.1016/j.nbt.2019.01.005
PMID: 30659933

Postprint 30 p, 997.5 KB

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