Full humanization of the glycolytic pathway in Saccharomyces cerevisiae

Francine J. Boonekamp; Ewout Knibbe; Marcel A. Vieira-Lara; Melanie Wijsman; Marijke A.H. Luttik; Karen van Eunen; Maxime den Ridder; Reinier Bron; Ana Maria Almonacid Suarez; Patrick van Rijn; Justina C. Wolters; Martin Pabst; Jean Marc Daran; Barbara M. Bakker; Pascale Daran-Lapujade

doi:10.1016/j.celrep.2022.111010

Full humanization of the glycolytic pathway in Saccharomyces cerevisiae

Francine J. Boonekamp, Ewout Knibbe, Marcel A. Vieira-Lara, Melanie Wijsman, Marijke A.H. Luttik, Karen van Eunen, Maxime den Ridder, Reinier Bron, Ana Maria Almonacid Suarez, Patrick van Rijn, Justina C. Wolters, Martin Pabst, Jean Marc Daran, Barbara M. Bakker, Pascale Daran-Lapujade

Research output: Contribution to Journal › Article › Academic › peer-review

Abstract

Although transplantation of single genes in yeast plays a key role in elucidating gene functionality in metazoans, technical challenges hamper humanization of full pathways and processes. Empowered by advances in synthetic biology, this study demonstrates the feasibility and implementation of full humanization of glycolysis in yeast. Single gene and full pathway transplantation revealed the remarkable conservation of glycolytic and moonlighting functions and, combined with evolutionary strategies, brought to light context-dependent responses. Human hexokinase 1 and 2, but not 4, required mutations in their catalytic or allosteric sites for functionality in yeast, whereas hexokinase 3 was unable to complement its yeast ortholog. Comparison with human tissues cultures showed preservation of turnover numbers of human glycolytic enzymes in yeast and human cell cultures. This demonstration of transplantation of an entire essential pathway paves the way for establishment of species-, tissue-, and disease-specific metazoan models.

Original language	English
Article number	111010
Number of pages	1
Journal	Cell Reports
Volume	39
Issue number	13
DOIs	https://doi.org/10.1016/j.celrep.2022.111010
Publication status	Published - 28 Jun 2022
Externally published	Yes

Bibliographical note

Publisher Copyright:
Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.

Funding

We thank Jordi Geelhoed for reverse engineering and growth characterization, Agnes Hol and Ingeborg van Lakwijk for valuable work on the human hexokinase strains, and Lycka Kamoen, Eveline Vreeburg, and Daniel Solis-Escalante for contributions to construction of reference strains. We thank Carol de Ram for processing proteomics samples, Pilar de la Torre and Marcel van den Broek for whole-genome sequencing and analysis, and Philip de Groot for help with bioreactors. This project was funded by the AdLibYeast European Research Council consolidator 648141 grant (to P.D.-L.), a UMCG-GSMS PhD fellowship, and the De Cock-Hadders Foundation .

Funders	Funder number
AdLibYeast European Research Council	648141
Stichting De Cock-Hadders

Keywords

adaptive laboratory evolution
CP: Metabolism
glycolysis
hexokinase
humanized yeast model
moonlighting function
pathway transplantation
Saccharomyces cerevisiae
synthetic biology

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Boonekamp, F. J., Knibbe, E., Vieira-Lara, M. A., Wijsman, M., Luttik, M. A. H., van Eunen, K., Ridder, M. D., Bron, R., Almonacid Suarez, A. M., van Rijn, P., Wolters, J. C., Pabst, M., Daran, J. M., Bakker, B. M., & Daran-Lapujade, P. (2022). Full humanization of the glycolytic pathway in Saccharomyces cerevisiae. Cell Reports, 39(13), Article 111010. https://doi.org/10.1016/j.celrep.2022.111010

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Full humanization of the glycolytic pathway in Saccharomyces cerevisiae

Abstract

Bibliographical note

Funding

Keywords

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