High pressure homogenization is a key unit operation in inclusion body processing

Britta Eggenreich; David Johannes Wurm; Vignesh Rajamanickam; Robert Klausser; Christoph Slouka; Oliver Spadiut

doi:10.1016/j.btecx.2020.100022

High pressure homogenization is a key unit operation in inclusion body processing

J Biotechnol. 2020:324S:100022. doi: 10.1016/j.btecx.2020.100022. Epub 2020 Jun 30.

Authors

Britta Eggenreich¹, David Johannes Wurm², Vignesh Rajamanickam³, Robert Klausser⁴, Christoph Slouka⁵, Oliver Spadiut⁶

Affiliations

¹ Institute of Chemical, Environmental and Bioscience Engineering, Research Group Integrated Bioprocess Development, Technische Universität Wien, Gumpendorferstrasse 1a, 1060 Vienna, Austria. Electronic address: britta_eggenreich@hotmail.com.
² Institute of Chemical, Environmental and Bioscience Engineering, Research Group Integrated Bioprocess Development, Technische Universität Wien, Gumpendorferstrasse 1a, 1060 Vienna, Austria. Electronic address: david.wurm@tuwien.ac.at.
³ Institute of Chemical, Environmental and Bioscience Engineering, Research Group Integrated Bioprocess Development, Technische Universität Wien, Gumpendorferstrasse 1a, 1060 Vienna, Austria. Electronic address: vignesh.rajamanickam@tuwien.ac.at.
⁴ Institute of Chemical, Environmental and Bioscience Engineering, Research Group Integrated Bioprocess Development, Technische Universität Wien, Gumpendorferstrasse 1a, 1060 Vienna, Austria. Electronic address: e1225699@student.tuwien.ac.at.
⁵ Institute of Chemical, Environmental and Bioscience Engineering, Research Group Integrated Bioprocess Development, Technische Universität Wien, Gumpendorferstrasse 1a, 1060 Vienna, Austria. Electronic address: christoph.slouka@tuwien.ac.at.
⁶ Institute of Chemical, Environmental and Bioscience Engineering, Research Group Integrated Bioprocess Development, Technische Universität Wien, Gumpendorferstrasse 1a, 1060 Vienna, Austria. Electronic address: oliver.spadiut@tuwien.ac.at.

PMID: 34154734
DOI: 10.1016/j.btecx.2020.100022

Abstract

Recombinant protein production in E. coli often leads to the formation of inclusion bodies (IBs). Although downstream processing of IBs has the reputation of being a great hurdle, advantages of IBs can be substantial. Highly pure recombinant protein with the possibility of correctly folded structures and an easy separation from cell matter are decisive factors that make IB processes so interesting. Product yield, purity and biological activity of the refolded protein are the responses to evaluate an IB process. The objective of this case study was to develop a refolding process in an integrated manner. The effects of the unit operations 1) homogenization, 2) IB wash and 3) IB solubilisation as well as their interdependencies were analyzed. We revealed interesting factor interactions between homogenization and IB wash, as well as homogenization and solubilisation, which would be overlooked if the single unit operations were investigated individually. Furthermore, we found that homogenization was a key unit operation for IB processing. By changing the conditions during homogenization only, the product yield, purity and biological activity of the refolded product was affected 2-fold, 1.2-fold and 2.5-fold, respectively.

Keywords: High pressure homogenization; Inclusion body processing; Inclusion body size; Product purity; Refolding yield.

MeSH terms

Escherichia coli*
Inclusion Bodies*
Recombinant Proteins

Substances

Recombinant Proteins