Periplasmic Chaperones Used to Enhance Functional Secretion of Proteins in E. coli

Martin Schlapschy, Arne Skerra

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

40 Scopus citations


While Escherichia coli is in wide use as a host organism for preparative protein production, problems with the folding of the recombinant gene product as well as protein aggregation, i.e., formation of inclusion bodies, are frequently encountered. This is particularly true for proteins that carry structural disulfide bonds, including antibody fragments, cytokines, growth factors, and extracellular fragments of eukaryotic cell surface receptors. In these cases, secretion into the oxidizing milieu of the bacterial periplasm in principle enables disulfide bond formation, resulting in a correctly folded and soluble protein. However, this process often occurs at low efficiency, depending on the nature of the recombinant gene product. Therefore, we have developed the helper plasmid pTUM4, which effects overexpression of four established periplasmic chaperones and/or folding catalysts: the thiol-disulfide oxidoreductases DsbA and DsbC, which catalyze the formation and isomerization of disulfide bridges, and two peptidyl-prolyl cis/trans isomerases with chaperone activity, FkpA and SurA. Here, we present a detailed protocol how to use this system for the bacterial secretion of recombinant proteins, including human EGF as a new example, and we give hints on optimization of the expression procedure.

Original languageEnglish
Title of host publicationHeterologous Gene Expression in E.coli
Subtitle of host publicationMethods and Protocols
PublisherHumana Press Inc.
Number of pages14
ISBN (Print)9781617379666
StatePublished - 2011

Publication series

NameMethods in Molecular Biology
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029


  • Chaperone
  • DsbA
  • DsbC
  • FkpA
  • SurA
  • disulfide isomerase
  • folding catalyst
  • peptidyl-prolyl cis/trans isomerase


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