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Chaperones facilitate heterologous expression of naturally evolved putative de novo proteins

View ORCID ProfileLars A. Eicholt, View ORCID ProfileMargaux Aubel, View ORCID ProfileKatrin Berk, View ORCID ProfileErich Bornberg-Bauer, View ORCID ProfileAndreas Lange
doi: https://doi.org/10.1101/2022.03.02.482622
Lars A. Eicholt
1Institute for Evolution and Biodiversity, University of Muenster, Germany
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Margaux Aubel
1Institute for Evolution and Biodiversity, University of Muenster, Germany
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Katrin Berk
1Institute for Evolution and Biodiversity, University of Muenster, Germany
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Erich Bornberg-Bauer
1Institute for Evolution and Biodiversity, University of Muenster, Germany
2Max Planck-Institute for Biology Tuebingen, Germany
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Andreas Lange
1Institute for Evolution and Biodiversity, University of Muenster, Germany
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  • For correspondence: andreas.lange@wwu.de
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Abstract

Over the past decade, evidence has accumulated that new protein coding genes can emerge de novo from previously non-coding DNA. Most studies have focused on large scale computational predictions of de novo protein coding genes across a wide range of organisms. In contrast, experimental data concerning the folding and function of de novo proteins is scarce. This might be due to difficulties in handling de novo proteins in vitro, as most are predicted to be short and disordered. Here we propose a guideline for the effective expression of eukaryotic de novo proteins in Escherichia coli.

We used 11 sequences from Drosophila melanogaster and 10 from Homo sapiens, that are predicted de novo proteins from former studies, for heterologous expression. The candidate de novo proteins have varying secondary structure and disorder content. Using multiple combinations of purification tags, E. coli expression strains and chaperone systems, we were able to increase the number of solubly expressed putative de novo proteins from 30 % to 62 %. Our findings indicate that the best combination for expressing putative de novo proteins in E.coli is a GST-tag with T7 Express cells and co-expressed chaperones. We found that, overall, proteins with higher predicted disorder were easier to express.

Competing Interest Statement

The authors have declared no competing interest.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
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Posted March 02, 2022.
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Chaperones facilitate heterologous expression of naturally evolved putative de novo proteins
Lars A. Eicholt, Margaux Aubel, Katrin Berk, Erich Bornberg-Bauer, Andreas Lange
bioRxiv 2022.03.02.482622; doi: https://doi.org/10.1101/2022.03.02.482622
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Chaperones facilitate heterologous expression of naturally evolved putative de novo proteins
Lars A. Eicholt, Margaux Aubel, Katrin Berk, Erich Bornberg-Bauer, Andreas Lange
bioRxiv 2022.03.02.482622; doi: https://doi.org/10.1101/2022.03.02.482622

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