Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:Q07644 (polypeptide)
 72,197 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A membrane-bound cytochrome b, a heterodimer formed by a 91-kD glycoprotein (heavy chain) and a 22-kD polypeptide (light chain), is an essential component of the phagocyte NADPH-oxidase responsible for superoxide generation. Cytochrome b is absent in two subgroups of chronic granulomatous disease (CGD), an inherited disorder characterized by the lack of oxidase activity. Mutations in the cytochrome heavy chain gene, encoded by the CYBB locus in Xp21.1, result in the X-linked form of CGD. A rare subgroup of autosomal recessive CGD also lacks cytochrome b (A- CGD), but the genetic defect has not previously been identified. In order to search for possible mutations in the cytochrome light chain locus, CYBA, the structure of this gene was characterized. The CYBA locus was localized to 16q24, and the approximately 600-bp open reading frame determined to be encoded by six exons that span approximately 8.5 kb. Three unrelated patients with A- CGD were studied for evidence of mutations in the light chain gene. One patient, whose parents were first cousins, was homozygous for a large deletion that removed all but the extreme 5' coding sequence of the gene. The other two patients had a grossly normal light chain transcript on Northern blot of mononuclear cell RNA. The light chain transcript was amplified by the polymerase chain reaction and sequenced. One patient was a compound heterozygote for two alleles containing point mutations in the open reading frame that predict a frame shift and a nonconservative amino acid replacement, respectively. The second patient, whose parents were second cousins, was homozygous for a different single-base substitution resulting in another nonconservative amino acid change. These results indicate that A- CGD can results from defects in the gene encoding the 22-kD light chain of the phagocyte cytochrome b.
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PMID:Human neutrophil cytochrome b light chain (p22-phox). Gene structure, chromosomal location, and mutations in cytochrome-negative autosomal recessive chronic granulomatous disease. 224 41

The NADPH-linked 3-oxoacyl-(acyl-carrier protein) (ACP) reductase (EC 1.1.1.100), also known as 'beta-ketoacyl-ACP reductase', has been purified from the mesocarp of mature avocado pears (Persea americana). The enzyme is inactivated by low ionic strength and low temperature. On SDS/PAGE under reducing conditions, purified 3-oxoacyl-ACP reductase migrated as a single polypeptide giving a molecular mass of 28 kDa. Gel-filtration chromatography gave an apparent native molecular mass of 130 kDa, suggesting that the enzyme is tetrameric. The enzyme is inactivated by dilution, but some protection is afforded by the presence of NADPH. Kinetic constants have been determined using synthetic analogues as well as the natural ACP substrate. It exhibits a broad pH optimum around neutrality. Phenylglyoxal inactivates the enzyme, and partial protection is given by 1 mM-NADPH. Antibodies have been raised against the protein, which were used to localize it using immunogold electron microscopy. It is localized in plastids. N-Terminal amino-acid-sequence analysis was performed on the enzyme, and it shows close structural similarity with cytochrome f. Internal amino-acid-sequence data, derived from tryptic peptides, shows similarity with the putative gene products encoded by the nodG gene from the nitrogen-fixing bacterium Rhizobium meliloti and the gra III act III genes from Streptomyces spp.
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PMID:3-Oxoacyl-(acyl-carrier protein) reductase from avocado (Persea americana) fruit mesocarp. 224 75

Using fluorescence spectroscopy, we have demonstrated that isolated envelope membranes from mature spinach chloroplasts catalyze the phototransformation of endogenous protochlorophyllide into chlorophyllide in presence of NADPH, but not in presence of NADH. Protochlorophyllide reductase was characterized further using monospecific antibodies (anti-protochlorophyllide reductase) raised against the purified enzyme from oat. In mature spinach chloroplasts, protochlorophyllide reductase is present only in envelope membranes. We have demonstrated that the envelope protochlorophyllide reductase, a 37,000-dalton polypeptide, is only a minor envelope component and is present on the outer surface of the outer envelope membrane. This conclusion is supported by several lines of evidence: (a) the envelope polypeptide that was immunodecorated with anti-protochlorophyllide reductase can be distinguished from the major 37,000-dalton envelope polypeptide E37 (which was identified by monospecific antibodies) only after two-dimensional polyacrylamide gel electrophoresis; (b) the envelope protochlorophyllide reductase was hydrolyzed when isolated intact chloroplasts were incubated in presence of thermolysin; and (c) isolated intact chloroplasts strongly agglutinate when incubated in presence of antibodies raised against protochlorophyllide reductase. These results demonstrate that major differences exist between chloroplasts and etioplasts with respect to protochlorophyllide reductase levels and localization. The presence on the chloroplast envelope membrane of both the substrate (protochlorophyllide) and the enzyme (protochlorophyllide reductase) necessary for chlorophyllide synthesis could have major implications for the understanding of chlorophyll biosynthesis in mature chloroplasts.
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PMID:Envelope membranes from mature spinach chloroplasts contain a NADPH:protochlorophyllide reductase on the cytosolic side of the outer membrane. 225 34

An NADPH:2'-hydroxydaidzein oxidoreductase (HDR) from elicitor-challenged soybean cell cultures was purified to apparent homogeneity by a five-step procedure. The purification procedure included affinity adsorption on Blue Sepharose and elution of the enzyme with NADP+. It was shown by gel filtration and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis that HDR consists of only one polypeptide, which has a Mr about 34,700. The pH optimum of the reaction was 7.0. Apparent Michaelis constants determined for 2'-hydroxydaidzein, 2'-hydroxyformononetin, and NADPH were, respectively, 50, 60, and 56 microM. A low conversion of 2'-hydroxygenistein to the corresponding isoflavanone was also observed but isoflavones lacking a 2'-hydroxyl group and various other flavonoids did not serve as substrates. Enzymatically derived 2'-hydroxydihydrodaidzein gave a positive CD spectrum at 328 nm, which shows its 3R stereochemistry. Antibodies against HDR were raised in rats.
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PMID:Phytoalexin synthesis in soybean: purification and characterization of NADPH:2'-hydroxydaidzein oxidoreductase from elicitor-challenged soybean cell cultures. 230 2

Rifapentine (R773, DL473) is a long-acting antituberculous drug used in China. In our experiments we have found some manifestations of induction of hepatic mixed function oxidase system in mice following pretreatment with rifapentine or phenobarbital. Both rifapentine and phenobarbital significantly increased the rate of antipyrine and pentobarbital metabolism in vivo. They also increased liver weight, the content of liver microsomal protein and cytochrome P-450, the activity of NADPH-cytochrome C reductase and NADPH oxidase. SDS-polyacylamide gel electrophoresis showed that the relative proportions of some polypeptide bands in mice microsomal fraction were significantly changed following rifapentine or phenobarbital pretreatment. The results indicate that rifapentine, like phenobarbital, is a potent inducer of hepatic mixed function oxidase system in mice and that it should be used carefully in clinical therapy, when combined with other drugs.
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PMID:Inductive effects of rifapentine on mice hepatic mixed function oxidase system. 231 33

Cytochrome P-450BM-3 is a catalytically self-sufficient enzyme which monooxygenates saturated and unsaturated fatty acids, alcohols, and amides. The protein has two domains: one which contains heme and is P-450-like and the other which contains FAD and FMN and is P-450 reductase-like. Both domains are on a single polypeptide chain. Utilizing a plasmid containing the gene encoding P-450BM-3, we have transformed the Escherichia coli strain DH5 alpha. This clone overexpresses P-450BM-3 to make approximately 20% of the soluble protein of this organism under optimal conditions. P-450BM-3 can be purified to homogeneity from the soluble fraction of the protein of these cells with a recovery of 50% making this cell line an excellent source of this important enzyme. Purified preparations of P-450BM-3 hydroxylate palmitic acid at a rate of 1600 mol/min/mol of heme at 25 degrees C. The stoichiometry of NADPH to oxygen utilized was 1 for all conditions; however, the ratio of oxygen or NADPH utilized per molecule of fatty acid substrate metabolized was different for different homologs of saturated fatty acids, when low concentrations (less than 100 microM) of substrate were used. Lauric and myristic acids were metabolized to two hydroxylated products, irrespective of the initial concentration of fatty acid in the reaction mixture, and the ratio of oxygen consumed to fatty acid hydroxylated was 1. High concentrations of palmitic acid (greater than 200 microM) led to the formation of three polar metabolites and a stoichiometry of 1:1 was observed for oxygen and palmitic acid utilization. These results indicate that a single hydroxyl group was inserted into each of these molecules. Lower concentrations (less than 50 microM) of palmitic acid were metabolized to additional polar metabolites, and the ratio of oxygen consumed to fatty acid substrate consumed approximated 3:1. These results can be explained best by a hypothesis that the initial hydroxylated compounds, which accumulate during the oxidation of palmitic acid by P-450BM-3, can be further oxidized by this enzyme to polyhydroxy- or hydroxy-ketone products.
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PMID:Fatty acid monooxygenation by cytochrome P-450BM-3. 240 33

A soybean flour-induced, soluble cytochrome P-450 (P-450soy) was purified 130-fold to homogeneity from Streptomyces griseus. Native cytochrome P-450soy is a single polypeptide, with a molecular weight of 47,500, in association with one ferriprotoporphyrin IX prosthetic group. Oxidized P-450soy exhibited visible absorption maxima at 394, 514, and 646 nm, characteristic of a high-spin cytochrome P-450. The CO-reduced difference spectrum of P-450soy had a Soret maximum at 448 nm. When reconstituted with spinach ferredoxin and spinach ferredoxin:NADP+ oxidoreductase, purified cytochrome P-450soy catalyzed the NADPH-dependent oxidation of the xenobiotic substrates precocene II and 7-ethoxycoumarin. In vitro proteolysis of cytochrome P-450soy generated a stable and catalytically active cytochrome P-450, designated P-450soy delta.
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PMID:Purification and characterization of a soybean flour-induced cytochrome P-450 from Streptomyces griseus. 249 63

The monooxygenase, p-hydroxybenzoate hydroxylase (4-hydroxybenzoate, NADPH:oxygen oxidoreductase (3-hydroxylating), EC 1.14.13.2) has been isolated and purified from Pseudomonas aeruginosa. The reaction catalysed is linked to the pathways for degradation of aromatic compounds by microorganisms. The enzyme has been quantitatively characterized in this paper for use in the mechanistic analysis of the protein by site-directed mutagenesis. This can be achieved when the results presented are used in combination with the information on the sequence and structure of the gene for this protein and the high-resolution crystallographic data for the protein from P. fluorescens. The protein is a dimer of identical sub-units in solution, and has one FAD per polypeptide with a monomeric molecular weight of 45,000. A full steady-state kinetic analysis was carried out at the optimum pH (8.0). A Vmax of 3750 min-1 at 25 degrees C was calculated, and the enzyme has a concerted-substitution mechanism, involving the substrates, NADPH, oxygen, and p-hydroxybenzoate. Extensive analyses of the reactions of reduced enzyme with oxygen were carried out. The quality of the data obtained confirmed the mechanisms of these reactions as proposed earlier by the authors for the enzyme from P. fluorescens. It was found that the amino acid residue differences between enzyme from P. fluorescence and aeruginosa do marginally change some observed transient state kinetic parameters, even though the structure of the enzyme shows they have no direct role in catalysis. Thus, transient state kinetic analysis is an excellent tool to examine the role of amino acid residues in catalysis.
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PMID:Purification, properties, and oxygen reactivity of p-hydroxybenzoate hydroxylase from Pseudomonas aeruginosa. 251 88

A soluble cyclohexanone monooxygenase was purified 16.1-fold to homogeneity from a Xanthobacter sp. grown upon cyclohexane as sole source of carbon and energy. The native enzyme is a 50-kDa single polypeptide chain associated with FMN rather than FAD as flavin prosthetic group in a 1:1 stoichiometric relationship. The monooxygenase catalyses the transformation of cyclohexanone to the lactone 1-oxa-2-oxocycloheptane in an oxygen ring insertion reaction. Only related cycloalkanone substrates are accepted for oxygenation, no activity is shown towards straight-chain alkanones. Enzyme activity is strongly inhibited by sulphydryl-reactive agents, but is relatively insensitive to metal chelators, electron transport inhibitors and the metal ions Fe3+ and Cu2+. Cyclohexanone monooxygenase has Km values for cyclohexanone and NADPH of less than 0.5 microM and 12.5 microM respectively. Kinetic investigations under steady-state conditions demonstrate that the flavoprotein prosthetic group, FMN, is involved in the monooxygenase catalytic mechanism. The systematic name for the enzyme is cyclohexanone, NADPH:oxygen oxidoreductase (6-hydroxylating, 1,2-lactonizing) (EC 1.14.13.22).
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PMID:Characterization of an FMN-containing cyclohexanone monooxygenase from a cyclohexane-grown Xanthobacter sp. 254 Sep 66

Cytochrome P-450BM-3 (P-450BM-3) from Bacillus megaterium incorporates both a P-450 and an NADPH:P-450 reductase in proteolytically separable domains of a single, 119-kDa polypeptide and functions as a fatty acid monooxygenase independently of any other protein. A 5-kilobase DNA fragment which contains the gene encoding P-450BM-3 was sequenced. A single continuous open reading frame starting at nucleotide 1541 of the 5-kilobase fragment correctly predicted the previously determined NH2-terminal protein sequences of the trypsin-generated P-450 and reductase domains and, in toto, predicted a mature polypeptide of 1,048-amino acid residues with Mr = 117,641. The trypsin site was found at arginine residue 471. The P-450 domain is most similar (about 25%) to the fatty acid omega-hydroxylases of P-450 family IV, while the reductase domain exhibits some 33% sequence similarity with the NADPH:P-450 reductases of mammalian liver. Both the P-450 and reductase domains of P-450BM-3 define new gene families but contain highly conserved segments which display as much as 50% sequence similarity with P-450s and reductases of eukaryotic origin. The mRNA for P-450BM-3 was found by S1 mapping to be 3,339 +/- 10 nucleotides in length. In the accompanying paper, two regions in the 1.5 kilobases 5' to the P-450BM-3 coding region have been implicated in the regulation of P-450BM-3 gene expression.
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PMID:Coding nucleotide, 5' regulatory, and deduced amino acid sequences of P-450BM-3, a single peptide cytochrome P-450:NADPH-P-450 reductase from Bacillus megaterium. 254 78


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