Expression of fungal MN peroxidase in E. coli, and refolding to yield active enzyme

Rose E. Whitwam, Irina G. Gazarian, Ming Tien

Research output: Contribution to journalArticle

50 Scopus citations

Abstract

The cDNA encoding Mn peroxidase isozyme H4 from Phanerochoete chrysosporium was expressed in Escherichia coli. The portion of the cDNA encoding the enzyme's signal peptide, not found in the processed holoenzyme, was deleted from the cDNA. The polypeptide was produced as inactive inclusion bodies that could be solubilized in 8 M urea and the reducing agent dithiothreitol. Reconstitution of activity was accomplished by diluting the urea concentration to 2M in the presence of hemin, calcium, and oxidized glutathione. All of the additives were required for recovery of activity. The activity of the recombinant enzyme was dependent on both Mn2+ and H202.

Original languageEnglish (US)
Pages (from-to)1013-1017
Number of pages5
JournalBiochemical and Biophysical Research Communications
Volume216
Issue number3
DOIs
StatePublished - Nov 22 1995

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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