Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.11.1.7 (peroxidase)
65,474 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Limited tryptic digestion of fluorescein isothiocyanate (FITC)-labeled (H+-K+)-ATPase from rat resting light gastric membranes produced a soluble 27-kDa polypeptide which retained the fluorescence of the parent enzyme. Its production was markedly enhanced in the presence of an amphiphilic detergent, Zwittergent 3-14, which potently inhibits the ATPase activity. This increase is probably due to protection of certain tryptic cleavage sites through conformational changes of the membrane enzyme by the detergent. The NH2-terminal sequence of the 27-kDa polypeptide corresponded exactly to that beginning at Asn-369 of the cDNA-deduced primary structure of the rat ATPase. The presence of the phosphorylation site, Asp-385, and FITC-labeled Lys-517, which is known to be a part of the ATP-binding site, indicates that the 27-kDa polypeptide contains a major cytoplasmic portion of (H+-K+)-ATPase. Interestingly, the polypeptide was stained with periodate-Schiff's base, indicating its glycoprotein nature. The carbohydrate group attached to the polypeptide seems to include at least an N-linked high-mannose moiety, since the polypeptide showed Con A binding activity as detected with a Con A-biotin/avidin-peroxidase assay on nitrocellulose transblots. Also, its Con A binding activity was inhibited by excess methyl alpha-D-mannopyranoside and disappeared upon treatment of the polypeptide with endoglycosidase H and N-glycanase. Further tryptic action converted the 27-kDa polypeptide to 2 smaller FITC-labeled polypeptides of 25 and 15 kDa, which lost 18 and 96 amino acid residues, respectively, from the NH2 terminus of the parent polypeptide.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Evidence for the presence of a carbohydrate moiety in fluorescein isothiocyanate labeled fragments of rat gastric (H+-K+)-ATPase. 254 51

Using human myeloperoxidase cDNA as a probe, a chromosomal gene related to myeloperoxidase was isolated from a human gene library. Comparison of the amino acid sequence deduced from the nucleotide sequence of the cloned gene with that of human eosinophil peroxidase purified from buffy coats has indicated that the isolated gene is the chromosomal gene for human eosinophil peroxidase. Like human myeloperoxidase gene, human eosinophil peroxidase gene consists of 12 exons and 11 introns spanning about 12 kilobases. The gene can code for a protein of 715 amino acids with a calculated Mr of 81,036. The heavy chain and the light chain of eosinophil peroxidase were located on the COOH and NH2 terminus of the protein, respectively. The coding sequences of eosinophil peroxidase and myeloperoxidase show homologies of 72.4% at the nucleotide and 69.8% at the amino acid level, while little homology was found in the 5'-flanking region. Northern hybridization and S1 mapping analysis of RNA from human leukemic cells have indicated that the eosinophil peroxidase gene is expressed in the eosinophilic subline of human HL-60 cells but not in the neutrophilic subline or in parental HL-60 cells.
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PMID:Molecular cloning and characterization of a chromosomal gene for human eosinophil peroxidase. 255 Apr 61

alpha-Kallikrein was prepared using an improved purification protocol (Burger, D., Schleuning, W.-D. and Schapira, M. (1986) J. Biol. Chem. 261, 324-327) and was employed to reevaluate our previous observations indicating that kallikrein activates blood neutrophils by a mechanism requiring an uncleaved Mr 52,000 NH2-terminal heavy chain. Cellular activation was evaluated by measuring neutrophil aggregation, release of both vitamin B12 binding capacity and myeloperoxidase, and generation of superoxide anion. Whereas all these indicators were evoked by exposing neutrophils to f-Met-Leu-Phe, phorbol myristate acetate, zymosan activated serum, or the calcium ionophore A 23187, we show here that neutrophils incubated with alpha-kallikrein remained unactivated. Moreover, we demonstrate that this lack of activation is accompanied by an inability of neutrophils to specifically bind alpha-kallikrein.
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PMID:Studies on the human plasma kallikrein-kinin system: alpha-kallikrein does not directly activate blood neutrophils. 255 Oct 61

The mechanism of N-dealkylation by peroxidases of the Ca2+ indicator quin2 and analogs was investigated and compared with the mechanism of N-dealkylation of some N-methyl-substituted aromatic amines. Nitrogen-centered cation radicals were detected by ESR spectroscopy for all the compounds studied. Further oxidation of the nitrogen-centered cation radicals, however, was dependent upon the structure of the radical formed. In the case of quin2 and analogs, a carbon-centered radical could be detected using the spin trap 5,5-dimethyl-1-pyrroline N-oxide. By using the spin trap 2-methyl-2-nitrosopropane (tert-nitrosobutane), it was determined that the carbon-centered radical was formed due to loss of a carboxylic acid group. This indicated that bond breakage most likely occurred through a rearrangement reaction. Furthermore, extensive oxygen consumption was detected, which was in agreement with the formation of carbon-centered radicals, as they avidly react with molecular oxygen. Thus, reaction of the carbon-centered radical with oxygen most likely led to the formation of a peroxyl radical. The peroxyl radical decomposed into superoxide that was spin trapped by 5,5-dimethyl-1-pyrroline N-oxide and an unstable iminium cation. The iminium cation would subsequently hydrolyze to the monomethyl amine and formaldehyde. In the case of N-methyl-substituted aromatic amines, carbon-centered radicals were not detected during the peroxidase-catalyzed oxidation of these compounds. Thus, rearrangement of the nitrogen-centered radical did not occur. Furthermore, little or no oxygen consumption was detected, whereas formaldehyde was formed in all cases. These results indicated that the N-methyl-substituted amines were oxidized by a mechanism different from the mechanism found for quin2 and analogs.
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PMID:The oxidation of N-substituted aromatic amines by horseradish peroxidase. 255 33

We describe a new enzymic colorimetric method in which urea is measured in serum by use of a single reagent mixture. Ammonia produced by urea hydrolysis, catalyzed by urease, reacts with glutamate and ATP in the presence of glutamine synthetase. The ADP so produced is assayed in reactions catalyzed sequentially by pyruvate kinase and pyruvate oxidase in a system that generates hydrogen peroxide. The hydrogen peroxide is measured at 500 or 550 nm in a reaction catalyzed by horseradish peroxidase, with phenol/4-aminophenazone as the chromogen. The reaction is complete in 15 min at 37 degrees C. The standard curve is linear up to a urea concentration of 40 mmol/L. Precision is good; CVs ranged from 2.5% to 3.1%. Results by the present method compared well with those by a candidate Reference Method and are not subject to interferences from commonly used drugs and anticoagulants.
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PMID:Enzymic urea assay: a new colorimetric method based on hydrogen peroxide measurement. 256 17

The presence of gastrin-releasing peptide-like immunoreactivity in the rat brain was investigated by use of the indirect peroxidase-antiperoxidase technique. A high density of gastrin-releasing peptide-like immunoreactive terminals in the ventral pallidum, the interpenduncular nucleus and in substantia nigra, pars reticulata, was observed. Moreover, gastrin-releasing peptide-like immunoreactive perikarya were observed in the hypothalamic suprachiasmatic nucleus. Antisera raised against gastrin-releasing peptide have been shown to cross-react with substance P, another peptide highly concentrated in the substantia nigra, the ventral pallidum and the interpenduncular nucleus, and gastrin-releasing peptide-immunoreactivity in these areas has therefore been regarded as substance P immunoreactivity. To determine the antigenic epitope recognized by the antiserum raised against gastrin-releasing peptide, specificity studies were performed with known peptides fixed to nitrocellulose filter strips as well as preabsorptions with the same peptides on fixed brain sections containing the substantia nigra. From these experiments, it could be deduced that the antiserum recognizes an epitope within the peptide sequence: Val-Gly-His-Leu-Met-NH2. The antiserum cross-reacts with bombesin and alytesin, but not with substance P, allowing us to conclude that gastrin-releasing peptide or a very closely related peptide is present in areas of the rat central nervous system in which substance P has previously also been shown to be present.
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PMID:An immunohistochemical demonstration of gastrin-releasing peptide (GRP) in the rat substantia nigra. 260 12

A full length cDNA clone, pGTB38 (C. B. Pickett et al. (1984) J. Biol. Chem. 259, 5182-5188), complementary to a rat liver glutathione S-transferase Ya mRNA has been expressed in Escherichia coli. The cDNA insert was isolated from pGTB38 using MaeI endonuclease digestion and was inserted into the expression vector pKK2.7 under the control of the tac promoter. Upon transformation of the expression vector into E. coli, two protein bands with molecular weights lower than the full-length Ya subunit were detected by Western blot analysis in the cell lysate of E. coli. These lower-molecular-weight proteins most likely result from incorrect initiation of translation at internal AUG codons instead of the first AUG codon of the mRNA. In order to eliminate the problem of incorrect initiation, the glutathione S-transferase Ya cDNA was isolated from the expression vector and digested with Bal31 to remove extra nucleotides from the 5' noncoding region. The protein expressed by this expression plasmid, pKK-GTB34, comigrated with the Ya subunit on sodium dodecyl sulfate polyacrylamide gels and was recognized by antibodies against the YaYc heterodimer. The expressed Ya homodimer was purified by S-hexylglutathione affinity and ion-exchange chromatographies. Approximately 50 mg pure protein was obtained from 9 liters of E. coli culture. The expressed Ya homodimer displayed glutathione-conjugating, peroxidase, and isomerase activities, which are identical to those of the native enzyme purified from rat liver cytosol. Protein sequencing indicates that the expressed protein has a serine as the NH2 terminus whereas the NH2 terminus of the glutathione S-transferase Ya homodimer purified from rat liver cytosol is apparently blocked.
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PMID:Expression of a cDNA encoding a rat liver glutathione S-transferase Ya subunit in Escherichia coli. 264 28

Myocardial ischemia is characterized by the liberation of adenosine and by complement-mediated inflammation. We have reported that amidated C3, formed when ammonia (NH3) disrupts the thiolester bond of C3, serves as an alternative pathway convertase, generates C5b-9, and stimulates phagocytic oxidative metabolism. We investigated whether the deamination of adenosine by adenosine deaminase in hematopoietic cells might liberate sufficient ammonia to form amidated C3 and thereby trigger complement-mediated inflammation at ischemic sites. In the presence of 4 mM adenosine, NH3 production per erythrocyte (RBC) was equal to that per neutrophil (PMN) (3.3 X 10(-15) mol/cell per h). Because RBC outnumber PMN in normal blood by a thousandfold, RBC are the major source of NH3 production in the presence of adenosine. NH3 production derived only from the deamination of adenosine by the enzyme adenosine deaminase and was abolished by 0.4 microM 2'-deoxycoformycin, a specific inhibitor of adenosine deaminase. When purified human C3 was incubated with 5 X 10(8) human RBC in the presence of adenosine, disruption of the C3 thiolester increased more than twofold over that measured in C3 incubated with buffer, or in C3 incubated with RBC (P less than 0.05). The formation of amidated C3 was abolished by the preincubation of RBC with 2'-deoxycoformycin (P less than 0.001). Amidated C3 elicited statistically significant release of superoxide, myeloperoxidase, and lactoferrin from PMN. Thus, the formation of amidated C3 by RBC deamination of adenosine triggers a cascade of complement-mediated inflammatory reactions.
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PMID:The erythrocyte as instigator of inflammation. Generation of amidated C3 by erythrocyte adenosine deaminase. 278 75

The heavy and the light subunits of human myeloperoxidase (donor: H2O2 oxidoreductase [EC 1.11.1.7]) I, II, and III were isolated from the reduced and S-carboxymethylated enzymes. These three enzymes have the same terminal amino acid sequences and similar chemical compositions in both subunits. The NH2-terminal sequences of the heavy and light subunits were determined to be Val-Asn-Cys-Glu-Thr- and Thr-Cys-Pro-Glu-Gln-, respectively; a heterogeneity was observed in the NH2-termini of the latter subunits for the three enzymes. As for COOH-termini, the sequences -(Asn, 2 Leu, Ala, Ser, Trp)-Arg-Glu-Ala and -Ala-Arg were obtained for the heavy and the light subunits, respectively. The heavy subunits contained 8-10 mol/mol of glucosamine. On the basis of these results and the amino acid sequence deduced from cDNA clones, the heavy subunits probably correspond to amino acids 279-744 and the light subunits to amino acids (164-167)-272. For the heavy subunits, Ser-745, which was predicted as the COOH-terminal amino acid from the nucleotide sequence, was removed. The light subunits were also processed at their COOH-termini by 6 residues. Four or five high mannose type carbohydrate chains were attached to the heavy subunits.
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PMID:Subunit structures of three human myeloperoxidases. 284 42

Somatostatin-28 (SS28) is considered as a precursor of somatostatin-14 (SS14) and also of the remaining NH2 terminus of SS28, the dodecapeptide SS28 which has been recently characterized. In order to study the cellular and subcellular localization of SS28 in the rat hypothalamus, we have conducted a light and electron microscopic immunocytochemical study, using both the peroxidase-antiperoxidase and the immunogold technique. Several antisera which selectively recognize one or more of these 3 somatostatin-related peptides were used. In serial paraffin sections, it was observed that SS28 was contained in the same neuronal cell bodies which also contain SS28. These neurons were exclusively located in the periventricular nucleus. All the antisera used also produced a strong staining in the external zone of the median eminence. At the electron microscopic level, the 3 peptides were exclusively localized in all the dense core vesicles in cell bodies and axons in the periventricular nucleus and terminals in the median eminence. These results strongly suggest that SS28 and SS14 originate from the precursor SS28 and that processing of the precursor occurs in the cell body. They also support the hypothesis that the 3 peptides are released simultaneously under appropriate stimulation.
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PMID:Immunocytochemical localization of somatostatin-28 in the rat hypothalamus. 285 89


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