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Integrated Autolysis, DNA Hydrolysis and Precipitation Enables an Improved Bioprocess for Q-Griffithsin, a Broad-Spectrum Antiviral and Clinical-Stage anti-COVID-19 Candidate

John S. Decker, Romel Menacho-Melgar, View ORCID ProfileMichael D. Lynch
doi: https://doi.org/10.1101/2021.12.30.474602
John S. Decker
1Department of Biomedical Engineering, Duke University, Durham, NC
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Romel Menacho-Melgar
1Department of Biomedical Engineering, Duke University, Durham, NC
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Michael D. Lynch
1Department of Biomedical Engineering, Duke University, Durham, NC
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  • ORCID record for Michael D. Lynch
  • For correspondence: michael.lynch@duke.edu
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Abstract

Across the biomanufacturing industry, innovations are needed to improve efficiency and flexibility, especially in the face of challenges such as the COVID-19 pandemic. Here we report an improved bioprocess for Q-Griffithsin, a broad-spectrum antiviral currently in clinical trials for COVID-19. Q-Griffithsin is produced at high titer in E. coli and purified to anticipated clinical grade without conventional chromatography or the need for any fixed downstream equipment. The process is thus both low-cost and highly flexible, facilitating low sales prices and agile modifications of production capacity, two key features for pandemic response. The simplicity of this process is enabled by a novel unit operation that integrates cellular autolysis, autohydrolysis of nucleic acids, and contaminant precipitation, giving essentially complete removal of host cell DNA as well as reducing host cell proteins and endotoxin by 3.6 and 2.4 log10 units, respectively. This unit operation can be performed rapidly and in the fermentation vessel, such that Q-GRFT is obtained with 100% yield and >99.9% purity immediately after fermentation and requires only a flow-through membrane chromatography step for further contaminant removal. Using this operation or variations of it may enable improved bioprocesses for a range of other high-value proteins in E. coli.

Highlights

  • Integrating autolysis, DNA hydrolysis and precipitation enables process simplification

  • Autolysis reduces endotoxin release and burden to purification

  • Q-Griffithsin recovered from fermentation vessel at >99.9% purity and 100% yield

  • Q-Griffithsin purified to anticipated clinical grade without conventional chromatography

  • The resulting bioprocess is 100% disposables-compatible, scalable, and low-cost

Competing Interest Statement

JSD, RMM and MDL have financial interests in Roke Biotechnologies, LLC. MDL has a financial interest in DMC Biotechnologies, Inc.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. All rights reserved. No reuse allowed without permission.
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Posted January 03, 2022.
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Integrated Autolysis, DNA Hydrolysis and Precipitation Enables an Improved Bioprocess for Q-Griffithsin, a Broad-Spectrum Antiviral and Clinical-Stage anti-COVID-19 Candidate
John S. Decker, Romel Menacho-Melgar, Michael D. Lynch
bioRxiv 2021.12.30.474602; doi: https://doi.org/10.1101/2021.12.30.474602
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Integrated Autolysis, DNA Hydrolysis and Precipitation Enables an Improved Bioprocess for Q-Griffithsin, a Broad-Spectrum Antiviral and Clinical-Stage anti-COVID-19 Candidate
John S. Decker, Romel Menacho-Melgar, Michael D. Lynch
bioRxiv 2021.12.30.474602; doi: https://doi.org/10.1101/2021.12.30.474602

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