Towards a generic prototyping approach for therapeutically-relevant peptides and proteins in a cell-free translation system

Yue Wu; Zhenling Cui; Yen-Hua Huang; Simon J de Veer; Andrey V Aralov; Zhong Guo; Shayli V Moradi; Alexandra O Hinton; Jennifer R Deuis; Shaodong Guo; Kai-En Chen; Brett M Collins; Irina Vetter; Volker Herzig; Alun Jones; Matthew A Cooper; Glenn F King; David J Craik; Kirill Alexandrov; Sergey Mureev

doi:10.1038/s41467-021-27854-9

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Towards a generic prototyping approach for therapeutically-relevant peptides and proteins in a cell-free translation system

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Towards a generic prototyping approach for therapeutically-relevant peptides and proteins in a cell-free translation system

Yue Wu, Zhenling Cui, Yen-Hua Huang, Simon J de Veer, Andrey V Aralov, Zhong Guo, Shayli V Moradi, Alexandra O Hinton, Jennifer R Deuis, Shaodong Guo, …

Nature Communications, Vol.13(1), pp.1-17

2022

DOI: https://doi.org/10.1038/s41467-021-27854-9

PMCID: PMC8752827

PMID: 35017494

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Abstract

Synthetic biology

Advances in peptide and protein therapeutics increased the need for rapid and cost-effective polypeptide prototyping. While in vitro translation systems are well suited for fast and multiplexed polypeptide prototyping, they suffer from misfolding, aggregation and disulfide-bond scrambling of the translated products. Here we propose that efficient folding of in vitro produced disulfide-rich peptides and proteins can be achieved if performed in an aggregation-free and thermodynamically controlled folding environment. To this end, we modify an E. coli-based in vitro translation system to allow co-translational capture of translated products by affinity matrix. This process reduces protein aggregation and enables productive oxidative folding and recycling of misfolded states under thermodynamic control. In this study we show that the developed approach is likely to be generally applicable for prototyping of a wide variety of disulfide-constrained peptides, macrocyclic peptides with non-native bonds and antibody fragments in amounts sufficient for interaction analysis and biological activity assessment.

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Title: Towards a generic prototyping approach for therapeutically-relevant peptides and proteins in a cell-free translation system
Authors: Yue Wu (Author) - The University of Queensland
Zhenling Cui (Author) - Queensland University of Technology
Yen-Hua Huang (Author) - The University of Queensland
Simon J de Veer (Author) - The University of Queensland
Andrey V Aralov (Author) - Siberian Branch of the Russian Academy of Sciences
Zhong Guo (Author) - Centre for Genomics and Personalised Health, Queensland University of Technology, Brisbane, QLD, 4001, Australia
Shayli V Moradi (Author) - Queensland University of Technology
Alexandra O Hinton (Author) - The University of Queensland
Jennifer R Deuis (Author) - Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD, 4072, Australia
Shaodong Guo (Author) - The University of Queensland
Kai-En Chen (Author) - The University of Queensland
Brett M Collins (Author) - The University of Queensland
Irina Vetter (Author) - The University of Queensland
Volker Herzig (Author) - University of the Sunshine Coast, Queensland, School of Science, Technology and Engineering
Alun Jones (Author) - Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD, 4072, Australia
Matthew A Cooper (Author) - The University of Queensland
Glenn F King (Author) - The University of Queensland
David J Craik (Author) - ARC Centre of Excellence for Innovations in Peptide and Protein Science
Kirill Alexandrov (Author) - Queensland University of Technology
Sergey Mureev (Author) - Queensland University of Technology
Publication details: Nature Communications, Vol.13(1), pp.1-17
Publisher: Nature Publishing Group
Date published: 2022
DOI: 10.1038/s41467-021-27854-9
ISSN: 2041-1723
PMID: 35017494; PMC8752827
Copyright note: © The Author(s) 2022. 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 license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license 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 license, visit http://creativecommons.org/licenses/by/4.0/.
Organisation Unit: University of the Sunshine Coast, Queensland; School of Science, Technology and Engineering; Centre for Bioinnovation
Language: English
Record Identifier: 99603407002621
Output Type: Journal article

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Collaboration types: Domestic collaboration; International collaboration
Web Of Science research areas: Biochemistry & Molecular Biology

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