Co-expression of ferrochelatase allows for complete heme incorporation into recombinant proteins produced in E. coli

Jawahar Sudhamsu, Mariam Kabir, Michael V. Airola, Bhumit A. Patel, Syun Ru Yeh, Denis L. Rousseau, Brian R. Crane

Research output: Contribution to journalArticle

19 Scopus citations

Abstract

Over-expression of heme binding proteins in Escherichia coli often results in sub-optimal heme incorporation and the amount of heme-bound protein produced usually varies with the protein of interest. Complete heme incorporation is important for biochemical characterization, spectroscopy, structural studies, and for the production of homogeneous commercial proteins with high activity. We have determined that recombinant proteins expressed in E. coli often contain less than a full complement of heme because they rather are partially incorporated with free-base porphyrin. Porphyrin-incorporated proteins have similar spectral characteristics as the desired heme-loaded targets, and thus are difficult to detect, even in purified samples. We present a straightforward and inexpensive solution to this problem that involves the coexpression of native ferrochelatase with the protein of interest. The method is shown to be effective for proteins that contain either Cys- or His-ligated hemes.

Original languageEnglish (US)
Pages (from-to)78-82
Number of pages5
JournalProtein Expression and Purification
Volume73
Issue number1
DOIs
StatePublished - 2010

Keywords

  • Ferrochelatase
  • Free-base porphyrin
  • Heme incorporation
  • Heme proteins

ASJC Scopus subject areas

  • Biotechnology

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