Protein purification from protein aggregates

Research output: Chapter in BookChapterResearchpeer-review

Abstract

© 2018 Nova Science Publishers, Inc. All rights reserved. Many recombinant proteins are nowadays produced in Escherichia coli. However, the production of these recombinant proteins in their soluble form is not an easy task. The aggregation of these proteins forming insoluble protein aggregates, known as inclusion bodies (IBs), is one of the most important bottlenecks in the production process. In these cases it has been necessary to develop protocols for the isolation of soluble recombinant proteins from IBs, which can be easily produced and isolated. Traditionally, based on the premise that IBs are formed by inactive proteins, these protocols have been based in three important steps: i) bacterial cell disruption and isolation of IBs; ii) IB solubilization with strong denaturants (high concentrations of chaotropic agents); iii) protein refolding and soluble protein purification. However, since in the last decade it has been proven that IBs are biologically active and structurally organized protein aggregates, new and adapted protocols have been developed to release the soluble protein retained in the IB scaffold without the need to use complex denaturing-refolding procedures. This new generation of protocols is based on a simple and mild solubilization process to preserve the existing structure and activity of IB-forming proteins. This chapter will detail different mild protocols, including those based on low concentrations of organic solvents or detergents, the modification of pH, and the application of high hydrostatic pressure or different cycles of freeze-thawing. All of them aimed to establish alternative protocols to traditional ones to isolate functional soluble proteins through an easily applicable protocol.
Original languageEnglish
Title of host publicationHandbook on Protein Purification: Industry Challenges and Technological Developments
Pages79-104
Number of pages25
Publication statusPublished - 15 Nov 2018

Keywords

  • Escherichia coli
  • Inclusion bodies
  • Mild solubilization
  • Recombinant proteins
  • Soluble proteins

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