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

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    16 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 - 26 Aug 2013

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