Crystallization of recombinant human heme oxygenase-1

Heme oxygenase catalyzes the NADPH, O-2 and cytochrome P450 reductase depen dent oxidation of heme to biliverdin and carbon monoxide. One of two primar y isozymes, HO-1, is anchored to the endoplasmic reticulum membrane via a s tretch of hydrophobic residues at the C-terminus. While full-length human H O-I consists of 288 residues, a truncated version with residues 1-265 has b een expressed as a soluble active enzyme in Escherichia coli. The recombina nt enzyme crystallized from ammonium sulfate solutions but the crystals wer e not of sufficient quality for diffraction studies. SDS gel analysis indic ated that the protein had undergone proteolytic degradation An increase in the use of protease inhibitors during purification eliminated proteolysis, but the intact protein did not crystallize. N-terminal sequencing and mass spectral analysis of dissolved crystals indicated that thr protein had degr aded to two major species consisting of residues 1-226 and 1-237. Expressio n of the 1-226 and 1-233 versions of human HO-1 provided active enzyme that crystallizes in a form suitable for diffraction studies. These crystals be long to space group P2(1), with unit cell dimensions a = 79.3 Angstrom, b = 56.3 Angstrom, c = 112.8 Angstrom, and beta = 101.5 degrees.