High Yield Production of a Soluble Human Interleukin-3 Variant from E. coli with Wild-Type Bioactivity and Improved Radiolabeling Properties

Timothy R. Hercus, Emma F. Barry, Mara Dottore, Barbara J. McClure, Andrew I. Webb, Angel F. Lopez, Ian G. Young, James M. Murphy

Research output: Contribution to journalArticlepeer-review

15 Citations (Scopus)

Abstract

Human interleukin-3 (hIL-3) is a polypeptide growth factor that regulates the proliferation, differentiation, survival and function of hematopoietic progenitors and many mature blood cell lineages. Although recombinant hIL-3 is a widely used laboratory reagent in hematology, standard methods for its preparation, including those employed by commercial suppliers, remain arduous owing to a reliance on refolding insoluble protein expressed in E. coli. In addition, wild-type hIL-3 is a poor substrate for radio-iodination, which has been a long-standing hindrance to its use in receptor binding assays. To overcome these problems, we developed a method for expression of hIL-3 in E. coli as a soluble protein, with typical yields of >3mg of purified hIL-3 per litre of shaking microbial culture. Additionally, we introduced a non-native tyrosine residue into our hIL-3 analog, which allowed radio-iodination to high specific activities for receptor binding studies whilst not compromising bioactivity. The method presented herein provides a cost-effective and convenient route to milligram quantities of a hIL-3 analog with wild-type bioactivity that, unlike wild-type hIL-3, can be efficiently radio-iodinated for receptor binding studies.

Original languageEnglish
Article numbere74376
JournalPloS one
Volume8
Issue number8
DOIs
Publication statusPublished or Issued - 26 Aug 2013
Externally publishedYes

ASJC Scopus subject areas

  • Agricultural and Biological Sciences(all)
  • General
  • Biochemistry, Genetics and Molecular Biology(all)

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