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Please use this identifier to cite or link to this item: https://doi.org/10.21256/zhaw-19480
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dc.contributor.authorRaschmanová, Hana-
dc.contributor.authorZamora, Iwo-
dc.contributor.authorBorčinová, Martina-
dc.contributor.authorMeier, Patrick-
dc.contributor.authorWeninger, Astrid-
dc.contributor.authorMächler, Dominik-
dc.contributor.authorGlieder, Anton-
dc.contributor.authorMelzoch, Karel-
dc.contributor.authorKnejzlík, Zdeněk-
dc.contributor.authorKovar, Karin-
dc.date.accessioned2020-02-19T14:14:53Z-
dc.date.available2020-02-19T14:14:53Z-
dc.date.issued2019-
dc.identifier.issn1664-302Xde_CH
dc.identifier.urihttps://digitalcollection.zhaw.ch/handle/11475/19480-
dc.description.abstractPichia pastoris (Komagataella sp.) is broadly used for the production of secreted recombinant proteins. Due to the high rate of protein production, incorrectly folded proteins may accumulate in the endoplasmic reticulum (ER). To restore their proper folding, the cell triggers the unfolded protein response (UPR); however, if the proteins cannot be repaired, they are degraded, which impairs process productivity. Moreover, a non-producing/non-secreting subpopulation of cells might occur, which also decreases overall productivity. Therefore, an in depth understanding of intracellular protein fluxes and population heterogeneity is needed to improve productivity. Under industrially relevant cultivation conditions in bioreactors, we cultured P. pastoris strains producing three different recombinant proteins: penicillin G acylase from Escherichia coli (EcPGA), lipase B from Candida antarctica (CaLB) and xylanase A from Thermomyces lanuginosus (TlXynA). Extracellular and intracellular product concentrations were determined, along with flow cytometry-based single-cell measurements of cell viability and the up-regulation of UPR. The cell population was distributed into four clusters, two of which were viable cells with no UPR up-regulation, differing in cell size and complexity. The other two clusters were cells with impaired viability, and cells with up-regulated UPR. Over the time course of cultivation, the distribution of the population into these four clusters changed. After 30 h of production, 60% of the cells producing EcPGA, which accumulated in the cells (50-70% of the product), had up-regulated UPR, but only 13% of the cells had impaired viability. A higher proportion of cells with decreased viability was observed in strains producing CaLB (20%) and TlXynA (27%). The proportion of cells with up-regulated UPR in CaLB-producing (35%) and TlXynA-producing (30%) strains was lower in comparison to the EcPGA-producing strain, and a smaller proportion of CaLB and TlXynA (<10%) accumulated in the cells. These data provide an insight into the development of heterogeneity in a recombinant P. pastoris population during a biotechnological process. A deeper understanding of the relationship between protein production/secretion and the regulation of the UPR might be utilized in bioprocess control and optimization with respect to secretion and population heterogeneity.de_CH
dc.language.isoende_CH
dc.publisherFrontiers Research Foundationde_CH
dc.relation.ispartofFrontiers in Microbiologyde_CH
dc.rightshttp://creativecommons.org/licenses/by/4.0/de_CH
dc.subjectPichia pastorisde_CH
dc.subjectFed-batch culturede_CH
dc.subjectFlow cytometryde_CH
dc.subjectHeterogeneityde_CH
dc.subjectSingle-cellde_CH
dc.subjectStress responsede_CH
dc.subjectSuper folder green fluorescent protein (sfGFP)de_CH
dc.subjectUnfolded protein response (UPR)de_CH
dc.subject.ddc660.6: Biotechnologiede_CH
dc.titleSingle-cell approach to monitor the unfolded protein response during biotechnological processes with pichia pastorisde_CH
dc.typeBeitrag in wissenschaftlicher Zeitschriftde_CH
dcterms.typeTextde_CH
zhaw.departementLife Sciences und Facility Managementde_CH
dc.identifier.doi10.3389/fmicb.2019.00335de_CH
dc.identifier.doi10.21256/zhaw-19480-
dc.identifier.pmid30873140de_CH
zhaw.funding.euNode_CH
zhaw.issue335de_CH
zhaw.originated.zhawYesde_CH
zhaw.publication.statuspublishedVersionde_CH
zhaw.volume10de_CH
zhaw.publication.reviewOpen peer reviewde_CH
zhaw.author.additionalNode_CH
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Raschmanová, H., Zamora, I., Borčinová, M., Meier, P., Weninger, A., Mächler, D., Glieder, A., Melzoch, K., Knejzlík, Z., & Kovar, K. (2019). Single-cell approach to monitor the unfolded protein response during biotechnological processes with pichia pastoris. Frontiers in Microbiology, 10(335). https://doi.org/10.3389/fmicb.2019.00335
Raschmanová, H. et al. (2019) ‘Single-cell approach to monitor the unfolded protein response during biotechnological processes with pichia pastoris’, Frontiers in Microbiology, 10(335). Available at: https://doi.org/10.3389/fmicb.2019.00335.
H. Raschmanová et al., “Single-cell approach to monitor the unfolded protein response during biotechnological processes with pichia pastoris,” Frontiers in Microbiology, vol. 10, no. 335, 2019, doi: 10.3389/fmicb.2019.00335.
RASCHMANOVÁ, Hana, Iwo ZAMORA, Martina BORČINOVÁ, Patrick MEIER, Astrid WENINGER, Dominik MÄCHLER, Anton GLIEDER, Karel MELZOCH, Zdeněk KNEJZLÍK und Karin KOVAR, 2019. Single-cell approach to monitor the unfolded protein response during biotechnological processes with pichia pastoris. Frontiers in Microbiology. 2019. Bd. 10, Nr. 335. DOI 10.3389/fmicb.2019.00335
Raschmanová, Hana, Iwo Zamora, Martina Borčinová, Patrick Meier, Astrid Weninger, Dominik Mächler, Anton Glieder, Karel Melzoch, Zdeněk Knejzlík, and Karin Kovar. 2019. “Single-Cell Approach to Monitor the Unfolded Protein Response during Biotechnological Processes with Pichia Pastoris.” Frontiers in Microbiology 10 (335). https://doi.org/10.3389/fmicb.2019.00335.
Raschmanová, Hana, et al. “Single-Cell Approach to Monitor the Unfolded Protein Response during Biotechnological Processes with Pichia Pastoris.” Frontiers in Microbiology, vol. 10, no. 335, 2019, https://doi.org/10.3389/fmicb.2019.00335.


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