Structural stability and heme binding potential of the truncated human dual oxidase 2 (DUOX2) peroxidase domain

Arch Biochem Biophys. 2011 Aug 15;512(2):197-203. doi: 10.1016/j.abb.2011.05.021. Epub 2011 Jun 17.

Abstract

The essential role of human dual oxidase 2 (hDUOX2) in thyroid hormone biosynthesis defines this member of the NOX/DUOX family, whose absence due to mutation has been directly related to disease, specifically hypothyroidism. Both human DUOX isoforms, hDUOX1 and hDUOX2, are expressed in thyroid tissue; however, hDUOX1 cannot compensate for inactivation of hDUOX2, suggesting that each enzyme is differentially regulated and/or functions in a unique manner. In efforts to uncover relevant structural and functional differences we have expressed and purified the peroxidase domain of hDUOX2(1-599) for direct comparison with the previously studied hDUOX1(1-593). As was shown for hDUOX1, the truncated hDUOX2 domain purifies without a bound heme co-factor and displays no peroxidase activity. However, hDUOX2(1-599) displays greater stability than hDUOX1(1-593). Surprisingly, upon titration with heme, both isoforms bind heme with a low micromolar affinity, demonstrating that they retain a heme binding site. A conformational difference in the full-length protein and/or a protein-protein interaction may be required to increase the heme binding affinity.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Amino Acid Sequence
  • Dual Oxidases
  • Enzyme Stability
  • Heme / metabolism
  • Humans
  • In Vitro Techniques
  • Models, Molecular
  • Molecular Sequence Data
  • NADPH Oxidases / chemistry*
  • NADPH Oxidases / genetics
  • NADPH Oxidases / metabolism*
  • Peptide Fragments / chemistry
  • Peptide Fragments / genetics
  • Peptide Fragments / metabolism
  • Protein Conformation
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Sequence Homology, Amino Acid
  • Solubility

Substances

  • Peptide Fragments
  • Recombinant Proteins
  • Heme
  • Dual Oxidases
  • NADPH Oxidases
  • DUOX1 protein, human
  • DUOX2 protein, human