Biochemical analysis of <em>Komagataella phaffii</em> oxidative folding proposes novel regulatory mechanisms of disulfide bond formation in yeast

Palma, Arianna; Rettenbacher, Lukas A.; Moilanen, Antti; Saaranen, Mirva; Pacheco-Martinez, Christian; Gasser, Brigitte; Ruddock, Lloyd

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Palma, A., Rettenbacher, L.A., Moilanen, A. et al. Biochemical analysis of Komagataella phaffii oxidative folding proposes novel regulatory mechanisms of disulfide bond formation in yeast. Sci Rep 13, 14298 (2023). https://doi.org/10.1038/s41598-023-41375-z

Biochemical analysis of Komagataella phaffii oxidative folding proposes novel regulatory mechanisms of disulfide bond formation in yeast

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Author: Palma, Arianna^1,2; Rettenbacher, Lukas A.^3,4; Moilanen, Antti⁴;
Saaranen, Mirva⁴; Pacheco-Martinez, Christian¹; Gasser, Brigitte^1,2; Ruddock, Lloyd⁴

Organizations: ¹Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna, Austria
²Austrian Centre of Industrial Biotechnology, Vienna, Austria
³School of Biosciences, University of Kent, Canterbury, UK

⁴Faculty of Biochemistry and Molecular Medicine, University of Oulu, Oulu, Finland

Format: article

Version: published version

Access: open

Online Access: PDF Full Text (PDF, 2.8 MB)

Persistent link: http://urn.fi/urn:nbn:fi-fe20231009139309

Language: English

Published: Springer Nature, 2023

Publish Date: 2023-10-09

Description:
Abstract

Oxidative protein folding in the endoplasmic reticulum (ER) is driven mainly by protein disulfide isomerase PDI and oxidoreductin Ero1. Their activity is tightly regulated and interconnected with the unfolded protein response (UPR). The mechanisms of disulfide bond formation have mainly been studied in human or in the yeast Saccharomyces cerevisiae. Here we analyze the kinetics of disulfide bond formation in the non-conventional yeast Komagataella phaffii, a common host for the production of recombinant secretory proteins. Surprisingly, we found significant differences with both the human and S. cerevisiae systems. Specifically, we report an inactive disulfide linked complex formed by K. phaffii Ero1 and Pdi1, similarly to the human orthologs, but not described in yeast before. Furthermore, we show how the interaction between K. phaffii Pdi1 and Ero1 is unaffected by the introduction of unfolded substrate into the system. This is drastically opposed to the previously observed behavior of the human pathway, suggesting a different regulation of the UPR and/or possibly different interaction mechanics between K. phaffii Pdi1 and Ero1.

see all


Series: Scientific reports

ISSN: 2045-2322

ISSN-E: 2045-2322

ISSN-L: 2045-2322

Volume: 13

Article number: 14298

DOI: 10.1038/s41598-023-41375-z

OADOI: https://oadoi.org/10.1038/s41598-023-41375-z

Type of Publication: A1 Journal article – refereed

Field of Science: 1182 Biochemistry, cell and molecular biology

Subjects:
Article

Funding: This work was funded by the Marie Skłodowska-Curie Actions Innovative Training Network of the European Union’s Horizon 2020 Program under grant agreement no. 813979 (SECRETERS). Further support was obtained by the Austrian Federal Ministry of Digital and Economic Affairs (BMDW), the Austrian Federal Ministry of Traffic, Innovation and Technology (bmvit), the Styrian Business Promotion Agency SFG, the Standortagentur Tirol, the Government of Lower Austria, the Business Agency Vienna and BOKU through the COMET Funding Program managed by the Austrian Research Promotion Agency FFG.

EU Grant Number: (813979) Secreters - A new generation of microbial expression hosts and tools for the production of biotherapeutics and high-value enzymes

Copyright information: © The Author(s) 2023. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

https://creativecommons.org/licenses/by/4.0/

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	Palma, A., Rettenbacher, L.A., Moilanen, A. et al. Biochemical analysis of Komagataella phaffii oxidative folding proposes novel regulatory mechanisms of disulfide bond formation in yeast. Sci Rep 13, 14298 (2023). https://doi.org/10.1038/s41598-023-41375-z Biochemical analysis of Komagataella phaffii oxidative folding proposes novel regulatory mechanisms of disulfide bond formation in yeast Saved in:
Author:	Palma, Arianna^1,2; Rettenbacher, Lukas A.^3,4; Moilanen, Antti⁴; Saaranen, Mirva⁴; Pacheco-Martinez, Christian¹; Gasser, Brigitte^1,2; Ruddock, Lloyd⁴
Organizations:	¹Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna, Austria ²Austrian Centre of Industrial Biotechnology, Vienna, Austria ³School of Biosciences, University of Kent, Canterbury, UK ⁴Faculty of Biochemistry and Molecular Medicine, University of Oulu, Oulu, Finland
Format:	article
Version:	published version
Access:	open
Online Access:	PDF Full Text (PDF, 2.8 MB)
Persistent link:	http://urn.fi/urn:nbn:fi-fe20231009139309
Language:	English
Published:	Springer Nature, 2023
Publish Date:	2023-10-09
Description:	Abstract Oxidative protein folding in the endoplasmic reticulum (ER) is driven mainly by protein disulfide isomerase PDI and oxidoreductin Ero1. Their activity is tightly regulated and interconnected with the unfolded protein response (UPR). The mechanisms of disulfide bond formation have mainly been studied in human or in the yeast Saccharomyces cerevisiae. Here we analyze the kinetics of disulfide bond formation in the non-conventional yeast Komagataella phaffii, a common host for the production of recombinant secretory proteins. Surprisingly, we found significant differences with both the human and S. cerevisiae systems. Specifically, we report an inactive disulfide linked complex formed by K. phaffii Ero1 and Pdi1, similarly to the human orthologs, but not described in yeast before. Furthermore, we show how the interaction between K. phaffii Pdi1 and Ero1 is unaffected by the introduction of unfolded substrate into the system. This is drastically opposed to the previously observed behavior of the human pathway, suggesting a different regulation of the UPR and/or possibly different interaction mechanics between K. phaffii Pdi1 and Ero1. see all 
Series:	Scientific reports
ISSN:	2045-2322
ISSN-E:	2045-2322
ISSN-L:	2045-2322
Volume:	13
Article number:	14298
DOI:	10.1038/s41598-023-41375-z
OADOI:	https://oadoi.org/10.1038/s41598-023-41375-z
Type of Publication:	A1 Journal article – refereed
Field of Science:	1182 Biochemistry, cell and molecular biology
Subjects:	Article
Funding:	This work was funded by the Marie Skłodowska-Curie Actions Innovative Training Network of the European Union’s Horizon 2020 Program under grant agreement no. 813979 (SECRETERS). Further support was obtained by the Austrian Federal Ministry of Digital and Economic Affairs (BMDW), the Austrian Federal Ministry of Traffic, Innovation and Technology (bmvit), the Styrian Business Promotion Agency SFG, the Standortagentur Tirol, the Government of Lower Austria, the Business Agency Vienna and BOKU through the COMET Funding Program managed by the Austrian Research Promotion Agency FFG.
EU Grant Number:	(813979) Secreters - A new generation of microbial expression hosts and tools for the production of biotherapeutics and high-value enzymes
Copyright information:	© The Author(s) 2023. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.
	https://creativecommons.org/licenses/by/4.0/