A screening strategy for heterologous protein expression in Escherichia coli with the highest return of investment

Protein Expr Purif. 2012 Jan;81(1):33-41. doi: 10.1016/j.pep.2011.08.030. Epub 2011 Sep 8.

Abstract

Heterologous protein expression in Escherichia coli is commonly used to obtain recombinant proteins for a variety of downstream applications. However, many proteins are not, or are only poorly, expressed in soluble form. High level expression often leads to the formation of inclusion bodies and an inactive product that needs to be refolded. By screening the solubility pattern for a set of 71 target proteins in different host-strains and varying parameters such as location of purification tag, promoter and induction temperature we propose a protocol with a success rate of 77% of clones returning a soluble protein. This protocol is particularly suitable for high-throughput screening with the goal to obtain soluble protein product for e.g. structure determination.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Chromatography, Affinity
  • Cloning, Molecular / methods*
  • Electrophoresis, Polyacrylamide Gel
  • Escherichia coli / genetics
  • Escherichia coli / metabolism*
  • Histidine
  • Oligopeptides
  • Recombinant Fusion Proteins / biosynthesis*
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Reproducibility of Results
  • Solubility
  • Temperature

Substances

  • His-His-His-His-His-His
  • Oligopeptides
  • Recombinant Fusion Proteins
  • Histidine