Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.14.99.3 (heme oxygenase)
 4,196 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The present report describes, for the first time, the identification of two constitutive forms of heme oxygenase, designated as HO-1 and HO-2, in rat liver microsomal fractions. HO-1 was purified to homogeneity and exhibited a specific activity of up to 4000 nmol of bilirubin/mg of protein/h. HO-2 was partially purified to a specific activity of 250 nmol of bilirubin/mg of protein/h. In the native state, the relative activity of HO-2 surpassed that of HO-1 by 2-3-fold. However, a remarkable difference existed in the regulatory mechanism(s) for the production of the two enzyme forms. Whereas the activity of HO-1 was increased up to 100-fold in response to cobalt, cadmium, hematin, phenylhydrazine, and bromobenzene, that of HO-2 was fully refractory to these agents. The two forms differed in their apparent Km, thermolability, ammonium sulfate precipitation, antigenicity, electrophoretic mobility under nondenaturing conditions, and chromatographic behavior. Specifically, for HO-1 the apparent Km value was 0.24 microM, whereas that for HO-2 was 0.67 microM. HO-2 preparation was more susceptible to heat inactivation; nearly 65% activity was retained by HO-1 preparation after exposure to 60 degrees C for 10 min, whereas under the same conditions only about 25% of HO-2 activity was retained. When subjected to ammonium sulfate precipitation the bulk of HO-1 activity precipitated between 0 and 35% saturation, whereas that of HO-2 was precipitated between 35 and 65% saturation. The two forms appeared as immunologically different entities, in so far as a crossreactivity between antibody raised against HO-1 in rabbit and HO-2 could not be detected. Similarities were observed in respect to cofactor requirements for activity, sensitivity to inhibitors, as well as their reactivity towards the substrates used in this study, i.e. hematin, hematoheme, and cytochrome c. Specifically both forms of the enzyme required NADPH-cytochrome c (P-450) reductase, NADPH or NADH, and O2 for activity, and reactions were inhibited by KCN, NaN3, and CO. Both forms cleaved the tetrapyrrole molecule exclusively at the alpha-meso bridge to form biliverdin IX alpha isomer. HO-1 and HO-2 utilized hematin and hematoheme as substrates but not intact cytochrome c.
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PMID:Characterization of two constitutive forms of rat liver microsomal heme oxygenase. Only one molecular species of the enzyme is inducible. 307 57

The concerted activity of two microsomal enzymes, heme oxygenase and NADPH-cytochrome c (P-450) reductase, is required for isomer-specific oxidation of heme molecule; heme oxygenase is commonly believed to be rate limiting in this activity. In this report, we provide evidence strongly suggesting the rate-limiting role of the reductase in oxidation of heme molecule in rat testis. In the testis and the liver of rats treated with Cd (20 mumol/kg, sc, 24 h) heme oxygenase activity, assessed by the formation of bilirubin, was decreased by 50% and increased by 7-fold, respectively. In these animals, the reductase activity was decreased by nearly 75% in the testis, but remained unchanged in the liver. Similarly, the reductase activity in the liver was not altered when heme oxygenase activity was increased by 20-fold in response to bromobenzene treatment. Addition of purified testicular reductase preparation (purified over 4000-fold), or hepatic reductase, to the testicular microsomes of Cd-treated rats obliterated the Cd-mediated inhibition of heme oxygenase activity. The chromatographic separation of heme oxygenase and the reductase of the testicular microsomal fractions revealed that the reductase activity was markedly decreased (75%) while the heme oxygenase activity, when assessed in the presence of exogenous reductase, was not affected by in vivo Cd treatment. In vitro, the membrane-bound reductase preparation obtained from the testis was more sensitive to the inhibitory effect of Cd than the liver preparation. However, the purified reductase preparations from the testis and the liver exhibited a similar degree of sensitivity to Cd. Based on the molar ratio of heme oxygenase to the reductase in the microsomal membranes of the liver and the testis it appeared that the testicular heme oxygenase, which is predominantly HO-2 isoform, interacts with the reductase less effectively than HO-1; in the induced liver, heme oxygenase is predominantly the HO-1 isoform. It is suggested that due to the low abundance of NADPH-cytochrome c (P-450) reductase and the apparently lower affinity of the enzyme for HO-2, the reductase exerts a regulatory action on heme oxygenase activity in the testis.
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PMID:Cadmium-mediated inhibition of testicular heme oxygenase activity: the role of NADPH-cytochrome c (P-450) reductase. 309 74

Two isoforms of heme oxygenase, designated as HO-1 and HO-2, were identified in rat spleen. The most abundant form was HO-1, wherein a relative ratio of about 5:1 of HO-1 to HO-2 was detected. The splenic HO-1 and HO-2 were immunochemically similar to the purified isoforms obtained from the liver and the testis. Moreover, the elution properties of splenic HO-1 as well as those of the constitutive liver HO-1 and the hematin-induced liver HO-1 on a DEAE-sephacel column were similar. However, the splenic HO-1 activity could not be induced by hematin. It is suggested that in the spleen heme oxygenase activity is maintained in the induced state as the result of constant exposure to hemoglobin released in the course of disruption of senescent erythrocytes.
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PMID:Characterization of two heme oxygenase isoforms in rat spleen: comparison with the hematin-induced and constitutive isoforms of the liver. 309 89

In this report we provide data, for the first time, demonstrating the conversion of the heme moiety of certain cytochrome P-450 and P-420 preparations, to biliverdin, catalyzed by heme oxygenase. We have used purified preparations of cytochromes P-450c, P-450b, P-450/P-420c, or P-450/P-420b as substrates in a heme oxygenase assay system reconstituted with heme oxygenase isoforms, HO-2 or HO-1, NADPH-cytochrome c (P-450) reductase, biliverdin reductase, NADPH, and Emulgen 911. With cytochrome P-450b or P-450/P-420b preparations, a near quantitative conversion of degraded heme to bile pigments was observed. In the case of cytochrome P-450/P-420c approximately 70% of the degraded heme was accounted for as bilirubin but only cytochrome P-420c was appreciably degraded. The role of heme oxygenase in this reaction was supported by the following observations: (i) bilirubin formation was not observed when heme oxygenase was omitted from the assay system; (ii) the rate of degradation of the heme moiety was at least threefold greater with heme oxygenase and NADPH-cytochrome c (P-450) reductase than that observed with reductase alone; and (iii) the presence of Zn- or Sn-protoporphyrins (2 microM), known competitive inhibitors of heme oxygenase, resulted in 70-90% inhibition of bilirubin formation.
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PMID:Rat liver cytochrome P-450b, P-420b, and P-420c are degraded to biliverdin by heme oxygenase. 312 60

In the present study we report on the detection of a distinct pattern of heme oxygenase isoform composition in the rat brain. In this organ only the noninducible form of heme oxygenase, HO-2, could be clearly detected. This pattern of composition distinguishes the brain from other organs tested to date, namely the liver, testis, and spleen. The rat brain microsomal fraction displayed a rather impressive rate of heme oxygenase activity. This fraction also exhibited a rate of NADPH-cytochrome P-450 reductase activity that was sufficient to fully support the oxygenase activity. The brain microsomal fraction was solubilized and subjected to ion-exchange chromatography on DEAE-Sephacel. The chromatographic elution pattern of heme oxygenase activity was compared with those of the liver and testis. In the brain only one peak of heme oxygenase activity was detected. The peak exhibited an elution profile similar to that of HO-2 of the liver and the testis. The presence of an activity peak was not detected in the elution profile at the region where the inducible isoform of heme oxygenase, HO-1, was expected. Cross-reactivity was observed between the solubilized brain microsomal fraction and antiserum to the testis HO-2 when subjected to Ouchterlony double diffusion immunoanalysis. A reaction was not observed when antiserum to liver HO-1 was employed. The presence of HO-2 in the brain microsomal preparation was also established by Western immunoblotting analysis. A protein having a mobility that was identical to the purified testicular HO-2 (Mr 36,000) was present in the brain microsomal preparation when probed with antiserum to HO-2. However, our attempts to demonstrate the presence of HO-1 in the brain microsomal preparation by a similar technique, but using antiserum to HO-1, were not successful. It is proposed that HO-2 is responsible for the bulk, if not all, of the brain microsomal heme oxygenase activity. It is further proposed that tissue-specific regulatory mechanisms are responsible for both the refractory response of the brain heme oxygenase to known metallic inducers and the absence of a detectable amount of the HO-1 isoform.
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PMID:Resolution of the rat brain heme oxygenase activity: absence of a detectable amount of the inducible form (HO-1). 312 61

In biological systems oxidation of heme is carried out by two isozymes of the microsomal heme oxygenase, HO-1 and HO-2. HO-1 is the commonly known heme oxygenase, the activity of which can be induced by up to 100-fold in response to a wide variety of stimuli (metals, heme, hormones, etc.). HO-2 was only recently discovered, and the isozyme appears to be uninducible. The two forms are products of two different genes and differ in their tissue expression. The primary structure of HO-1 and an HO-2 fragment of 91 amino acid residues show only 58% homology, but share a region with 100% secondary structure homology. This region is believed to be the catalytic site. Most likely, HO-1 gene is regulated in the same manner as metallothione in the gene. HO-1 has a heat shock regulatory element, and possibly many promoter elements, which bind to respective inducers and cause transcription of the gene. In vivo induction of HO-1 activity in the liver is accompanied by decreases in the total P-450 levels and, in a reconstituted system, cytochrome P-450b heme can be quantitatively converted to biliverdin by HO-1 and HO-2. The enzyme activity is inhibited in vivo for extended periods subsequent to binding of Zn- and Sn- protoporphyrins. This property appears useful for the suppression of bilirubin production. The metalloporphyrins, however, are not innocuous and cause major disruptions in cellular metabolism. In this review recent findings on heme oxygenase are highlighted.
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PMID:Heme oxygenase: function, multiplicity, regulatory mechanisms, and clinical applications. 329 25

Recently, we have reported on the presence of two forms of heme oxygenase in rat liver and testis microsomes, referred to as HO-1 and HO-2 (M. D. Maines, G. M. Trakshel, and R. K. Kutty (1986) J. Biol. Chem. 261, 411-419; G. M. Trakshel, R. K. Kutty, and M. D. Maines (1986) J. Biol. Chem. 261, 11131-11137). Although the two forms differed in several biochemical properties, we could not ascertain whether they represented two isozymes or whether they were isoforms of heme oxygenase. In the present study, we provide evidence suggesting that the two forms are isozymes and represent different gene products. We also provide data suggesting that HO-1 is the commonly known heme oxygenase form. The molecular weight and immunochemical properties of HO-1 and HO-2 did not vary depending on the tissue source examined, i.e. liver and testis. Major differences, however, were noted in the amino acid composition of the two forms including the presence of 3 cysteine/cystine residues in HO-2 only. Using antibody to HO-2, four testis clones and two liver clones were isolated, and one liver and one testis clone were sequenced. Both clones revealed a 274-base-pair insert, and the sequence of both inserts was the same. The validity of assignment was confirmed by matching a 14-amino-acid peptide obtained from purified HO-2 with the sequence. Approximately 43% amino acid homology was detected between the HO-2 insert and the published amino acid sequence of heme oxygenase. However, amino acid homology search revealed the presence of two regions of homology: one 22-mer sequence with only one unmatched amino acid, and one 10-mer sequence with one unmatched amino acid. Heme oxygenase appeared to be the HO-1 form, an assignment based on its amino acid sequence matching the sequence of 2 peptides obtained from purified HO-1 and the immunochemical properties of the cobalt-, hematin-, and bromobenzene-induced rat liver enzyme. The secondary structure prediction analysis revealed an area of 100% structural homology with only 72% sequence homology. We predict this region may represent the catalytic site of the enzyme.
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PMID:Evidence suggesting that the two forms of heme oxygenase are products of different genes. 334 48

This study shows heme oxygenase multiplicity is common to rat and human tissues. The isozymes in man and rat, however, are heterogenous proteins that share certain characteristics. Two forms of heme oxygenase, HO-1 and HO-2, were identified in human testis. HO-2 form was the prevalent form. Human and rat HO-1 differed in chromatographic behavior and molecular weight; human HO-1 was a larger molecule (35,400 vs 30,000). The two forms, however, were similar in that immunochemically human HO-1 exhibited reactivity toward antibody to rat HO-1. Human and rat HO-2 also were dissimilar in chromatographic behavior and showed only a weak immunological cross-reactivity. Human and rat HO-1 were essentially the same size. As in rat organs, the microsomal cytochrome P-450 content in human testis was reciprocal to heme oxygenase activity.
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PMID:Detection of two heme oxygenase isoforms in the human testis. 339 32

Recently we reported on the presence of two isoforms of heme oxygenase in rat liver microsomes, referred to as HO-1 and HO-2, and that only HO-1 is inducible (Maines, M. D., Trakshel, G. M., and Kutty, R. K. (1986) J. Biol. Chem. 261, 411-419). Presently we report on the detection of two isoforms of the enzyme in rat testis and purification to near homogeneity of the noninducible isoform, HO-2. A comparative characterization of the liver HO-1 and the testicular HO-2 is also provided. The relative abundance of the isoforms in the two organs was dissimilar. In the testis, the predominant form was HO-2, and only minute amounts of HO-1 were detected. In the liver, however, a 1:2 ratio of HO-1 to HO-2 was noted. The activity of HO-2 in both organs was refractory to cadmium, an inducer of the hepatic HO-1. Under nondenaturing electrophoresis conditions, HO-2 showed a higher mobility than HO-1; on a sodium dodecyl sulfate-polyacrylamide gel, HO-2 displayed a higher monomeric Mr. The apparent Mr values for HO-2 and HO-1 were 36,000 and 30,000, respectively. The isoforms differed in immunochemical properties. Antiserum to the liver HO-1 did not recognize the testicular HO-2 when examined by double immunodiffusion or by Western immunoblotting. HO-2 was more sensitive to heat inactivation than HO-1. When exposed at 65 degrees C (10 min), 70% of HO-1 activity was retained; however, nearly 80% of HO-2 activity was lost. The apparent Km values for heme for HO-1 and HO-2 were 0.24 and 0.40 microM, respectively. HO-1 and HO-2 had similar requirements for cofactor and flavoprotein reductase and were inhibited by heme-ligands (CO, KCN, NaN3). HO-2 utilized as substrate, Fe-protoporphyrin, Fe-hematoporphyrin, and Fe-hematoporphyrin acetate; it did not degrade intact purified rat liver cytochromes b5 and P-450 LM2, catalase, cytochrome c, hemoglobin, or myoglobin.
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PMID:Purification and characterization of the major constitutive form of testicular heme oxygenase. The noninducible isoform. 352 62

The distribution of the carbon monoxide (CO) producing enzymes haem oxygenase (HO) type 1 and 2 were studied in the feline lower oesophageal sphincter (LOS), as were HO activity and functional effects of CO. HO-2 immunoreactivity was observed in nerve cell bodies in the submucosal and myenteric plexus, nerve fibres, non-neuronal cells surrounding smooth muscle bundles, and in arterial endothelium, HO-1 immunoreactivity was confined to non-neuronal cells in the smooth muscle layer. CO production, indicating HO activity, was demonstrated in tissue homogenates. CO relaxed the LOS, and activated the cyclic GMP system. These results show that HO is present in the LOS, and suggest that CO can be generated by neuronal and non-neuronal structures and may have a role as a peripheral messenger.
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PMID:Carbon monoxide as a putative messenger molecule in the feline lower oesophageal sphincter of the cat. 748 31


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