Characterization of a cDNA-encoding rabbit brain heme oxygenase-2 and identification of a conserved domain among mammalian heme oxygenase isozymes: possible heme-binding site?
A 1.3-kb rat testis cDNA clone for heme oxygenase-2 (HO-2) was used as a Northern blot hybridization probe, and a single homologous mRNA species, of approximately 1.3 kb in rabbit brain and testis was detected. This contrasted with the observation made with rat brain in which two HO-2 transcripts of approximately 1.3 and 1.9 kb were detected. Use of the same rat HO-2 probe to screen a rabbit brain cDNA library in lambda gt11 resulted in the recovery of a single 1.2-kb cDNA clone. This cDNA exhibits 84% overall nucleotide sequence homology with rat HO-2 and encodes a protein of 35,352 Da, displaying 88% amino acid sequence homology with rat testis HO-2. Furthermore, when expressed in Escherichia coli, the rabbit cDNA-encoded protein displays heme oxygenase activity and cross-reactivity with antibody to rat HO-2. Based on findings obtained through Western immunoblot analysis of partially purified HO-2 protein prepared from rabbit testis and brain, the 35- to 36-kDa molecular form appears to be the major HO-2 form detected in the brain, whereas a 42-kDa species is the predominant form observed in rabbit testis. Having deduced the amino acid sequence of rabbit brain HO-2, we provide a comparison of this sequence with those of rat, mouse, and human HO-1 and rat HO-2, and thereby identify a 24-amino-acid-long peptide region which, except for one residue, is identical in all five species of HO-1 and HO-2 compared (96% similarity), and exhibits 100% similarity in predicted secondary structure (for this region) in all five proteins. We propose that this peptide may be important to the heme binding and isomer-specific tetrapyrrole cleavage activities of the heme oxygenase isozymes.
Department of Biophysics, University of Rochester School of Medicine, New York 14642.
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D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D001921:Brain / Q000201:enzymology*; D004247:DNA / Q000737:chemistry*; D006419:Heme Oxygenase (Decyclizing) / Q000235:genetics*; D006801:Humans; D007527:Isoenzymes / Q000235:genetics*; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011485:Protein Binding; D011487:Protein Conformation; D011817:Rabbits; D051381:Rats; D016415:Sequence Alignment; D012689:Sequence Homology, Nucleic Acid; D013737:Testis / Q000201:enzymology
D007527:Isoenzymes; D004247:DNA; D006419:Heme Oxygenase (Decyclizing)