Adeno-associated virus capsid protein expression in Escherichia coli and chemically defined capsid assembly

Le DT, Radukic M, Müller K (2019)
Scientific Reports 9: 18631.

Zeitschriftenaufsatz | Veröffentlicht| Englisch
 
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Abstract / Bemerkung
Research and clinical applications of recombinant adeno-associated virus (rAAV) significantly increased in recent years alongside regulatory approvals of rAAV gene therapy products. To date, all rAAV vectors as well as AAV empty capsids are produced in eukaryotic cells. We explored a new route to generate AAV capsids with the aim to analyze capsid assembly in a chemically defined setting and pave the way for new production methods and applications based on AAV virus-like particles (VLPs). We generated these empty capsids by bacterial expression and subsequent concomitant protein refolding and VLP formation. AAV serotype 2 structural protein VP3 was expressed in Escherichia coli. VLPs formed as demonstrated by dynamic light scattering, atomic force microscopy, and ELISA. Furthermore, VLPs internalized into human HeLa cells. To extend the application range of the VLPs, we tested peptide insertions, at the genetic level, in a surface loop (amino acid position 587) or at the C-terminus of VP3 and these variants also formed VLPs. VLPs developed without assembly-activating protein (AAP), but adding purified recombinant AAP to the refolding process increased capsid yield. Our findings offer a new route to understand AAV assembly biology and open a toolbox for AAV production strategies that might enable capsid display for vaccination and matching of capsids with cargoes at large scale and low cost.
Erscheinungsjahr
2019
Zeitschriftentitel
Scientific Reports
Band
9
Art.-Nr.
18631
eISSN
2045-2322
Finanzierungs-Informationen
Article Processing Charge funded by the Deutsche Forschungsgemeinschaft and the Open Access Publication Fund of Bielefeld University.
Page URI
https://pub.uni-bielefeld.de/record/2939510

Zitieren

Le DT, Radukic M, Müller K. Adeno-associated virus capsid protein expression in Escherichia coli and chemically defined capsid assembly. Scientific Reports. 2019;9: 18631.
Le, D. T., Radukic, M., & Müller, K. (2019). Adeno-associated virus capsid protein expression in Escherichia coli and chemically defined capsid assembly. Scientific Reports, 9, 18631. doi:10.1038/s41598-019-54928-y
Le, D. T., Radukic, M., and Müller, K. (2019). Adeno-associated virus capsid protein expression in Escherichia coli and chemically defined capsid assembly. Scientific Reports 9:18631.
Le, D.T., Radukic, M., & Müller, K., 2019. Adeno-associated virus capsid protein expression in Escherichia coli and chemically defined capsid assembly. Scientific Reports, 9: 18631.
D.T. Le, M. Radukic, and K. Müller, “Adeno-associated virus capsid protein expression in Escherichia coli and chemically defined capsid assembly”, Scientific Reports, vol. 9, 2019, : 18631.
Le, D.T., Radukic, M., Müller, K.: Adeno-associated virus capsid protein expression in Escherichia coli and chemically defined capsid assembly. Scientific Reports. 9, : 18631 (2019).
Le, Dinh To, Radukic, Marco, and Müller, Kristian. “Adeno-associated virus capsid protein expression in Escherichia coli and chemically defined capsid assembly”. Scientific Reports 9 (2019): 18631.
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