Heme oxygenase-2. Properties of the heme complex of the purified tryptic fragment of recombinant human heme oxygenase-2

J Biol Chem. 1995 Mar 17;270(11):6345-50. doi: 10.1074/jbc.270.11.6345.

Abstract

Recombinant human microsomal heme oxygenase-2 was expressed in Escherichia coli. Tryptic digestion of the membrane fraction, in which the wild-type enzyme was localized, yielded a soluble tryptic peptide of 28 kDa, which retained the ability to accept electrons from NADPH-cytochrome P-450 reductase and the enzymatic activity for conversion of heme to biliverdin. The tryptic fragment, when purified to apparent homogeneity, bound one equivalent of heme to form a substrate-enzyme complex that had spectroscopic properties characteristic of heme proteins, such as myoglobin and hemoglobin. Optical absorption, Raman scattering, and EPR studies of the heme-tryptic fragment complex revealed that the ferric heme was six coordinate high spin at neutral pH and six coordinate low spin at alkaline pH, with a pK alpha value of 8.5. EPR and Raman scattering studies indicated that a neutral imidazole of a histidine residue served as the proximal ligand in the heme-heme oxygenase-2 fragment complex. The reaction with hydrogen peroxide converted the heme of the heme oxygenase-2 fragment complex into a verdoheme-like intermediate, while the reaction with m-chloroperbenzoic acid yielded a oxoferryl species. These spectroscopic properties are similar to those obtained for heme oxygenase-1, and thus the catalytic mechanism of heme oxygenase-2 appears to be similar to that of heme oxygenase-1.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Cloning, Molecular
  • DNA, Complementary
  • Electron Spin Resonance Spectroscopy
  • Escherichia coli
  • Heme / chemistry
  • Heme / metabolism*
  • Heme Oxygenase (Decyclizing) / biosynthesis
  • Heme Oxygenase (Decyclizing) / chemistry*
  • Heme Oxygenase (Decyclizing) / metabolism*
  • Humans
  • Hydrogen-Ion Concentration
  • Isoenzymes / biosynthesis
  • Isoenzymes / chemistry
  • Isoenzymes / metabolism
  • Kinetics
  • Microsomes / enzymology
  • Molecular Sequence Data
  • Oligodeoxyribonucleotides
  • Peptide Fragments / chemistry
  • Peptide Fragments / metabolism*
  • Protein Conformation*
  • Rabbits
  • Rats
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism
  • Sequence Homology, Amino Acid
  • Spectrophotometry
  • Spectrum Analysis, Raman
  • Trypsin

Substances

  • DNA, Complementary
  • Isoenzymes
  • Oligodeoxyribonucleotides
  • Peptide Fragments
  • Recombinant Proteins
  • Heme
  • Heme Oxygenase (Decyclizing)
  • Trypsin

Associated data

  • GENBANK/D21243