Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Evidence showing that cytochrome P450-mediated reduction of brucine N-oxide to brucine by rat liver microsomes proceeds nonenzymatically in the presence of both a reduced pyridine nucleotide and FAD is presented. The microsomal N-oxide reduction appears to proceed in two steps: The first step is reduction of FAD by NADPH or NADH either enzymatically or nonenzymatically. The second step is nonenzymatic reduction of the tertiary amine N-oxide by the reduced flavin and is nonenzymatically catalyzed by the heme group of cytochrome P450.
Biochem Mol Biol Int 1997 Aug
PMID:Nonenzymatic reduction of brucine N-oxide by the heme group of cytochrome P450. 928 65

The cDNA fragments coding for the FAD-, glycerophosphate- and calcium-binding domains of mitochondrial glycerophosphate dehydrogenase (mGDH) were synthetized using RNA extracted from freshly isolated pancreatic islets of a normal subject and two-non-insulin-dependent diabetic patients. Single strand conformation polymorphism analysis of the PCR products yielded the same mobility as control cDNA probes. Likewise, the nucleotide sequence and corresponding amino acid sequence were identical to the normal gene bank sequence. These findings argue against the presence, in pancreatic islets, of an mGDH isoform distinct from that previously characterized in extrapancreatic organs.
Biochem Mol Biol Int 1997 Sep
PMID:Nucleotide sequence of cDNA fragments coding for the FAD-,glycerophosphate- and calcium-binding domains of human islet mitochondrial glycerophosphate dehydrogenase. 930 30

Thioredoxin reductase (TrxR) from Escherichia coli consists of two globular domains connected by a two-stranded beta sheet: an FAD domain and a pyridine nucleotide binding domain. The latter domain contains the redox-active disulfide composed of Cys 135 and Cys 138. TrxR is proposed to undergo a conformational change whereby the two domains rotate 66 degrees relative to each other (Waksman G, Krishna TSR, Williams CH Jr, Kuriyan J, 1994, J Mol Biol 236:800-816), placing either redox active disulfide (FO conformation) or the NADPH binding site (FR conformation) adjacent to the flavin. This domain rotation model was investigated by using a Cys 138 Ser active-site mutant. The flavin fluorescence of this mutant is only 7% that of wild-type TrxR, presumably due to the proximity of Ser 138 to the flavin in the FO conformation. Reaction of the remaining active-site thiol, Cys 135, with phenylmercuric acetate (PMA) causes a 9.5-fold increase in fluorescence. Titration of the PMA-treated mutant with the nonreducing NADP(H) analogue, 3-aminopyridine adenine dinucleotide phosphate (AADP+), results in significant quenching of the flavin fluorescence, which demonstrates binding adjacent to the FAD, as predicted for the FR conformation. Wild-type TrxR, with or without PMA treatment, shows similar quenching by AADP+, indicating that it exists mostly in the FR conformer. These findings, along with increased EndoGluC protease susceptibility of PMA-treated enzymes, agree with the model that the FO and FR conformations are in equilibrium. PMA treatment, because of steric limitations of the phenylmercuric adduct in the FO form, forces the equilibrium to the FR conformer, where AADP+ binding can cause fluorescence quenching and conformational restriction favors proteolytic susceptibility.
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PMID:Evidence for two conformational states of thioredoxin reductase from Escherichia coli: use of intrinsic and extrinsic quenchers of flavin fluorescence as probes to observe domain rotation. 933 41

The mitochondrial enzyme FAD-linked glycerophosphate dehydrogenase (mGDH) plays a key role in the recognition of glucose as a stimulus for insulin release from the pancreatic islet B-cell. In the present study, an ELISA procedure was used for the measurement of mGDH antibodies in both insulin-dependent (IDDM) and non-insulin-dependent (NIDDM) diabetic patients. Positive readings, exceeding the upper limit of the normal range, were recorded in 7 out of 12 IDDM patients, as distinct (P < 0.01) from 2 out of 12 nondiabetic subjects of comparable age. The study conducted in 41 NIDDM patients and 15 control subjects of similar age indicated that the incidence of mGDH-positive cases was not significantly different in the diabetic (4/41) and control (1/15) groups, the measurement of optical density in the positive cases barely exceeding the upper limit of the normal range. These findings indicate that the mitochondrial enzyme mGDH often acts as an antigenic determinant in IDDM, but not in NIDDM, patients.
Biochem Mol Med 1997 Dec
PMID:Enzyme-linked immunosorbent assay of autoantibodies against mitochondrial glycerophosphate dehydrogenase in insulin-dependent and non-insulin-dependent diabetic subjects. 944 69

Some general features of the respiratory chain and respiratory control were characterized in coupled mitochondrial preparations from Leishmania mexicana promastigotes. O2 uptake was sensitive to the electron-transfer inhibitors rotenone, flavone, malonate, 4,4,4-trifluoro-1-(2-thienyl) 1.3 butanedione (TTFA), antimycin A, 2n-nonyl-4-hydroxyquinoline-N-oxide (HQNO), myxothiazol, cyanide and azide. A high concentration of rotenone (60 microM) was required to inhibit O2 uptake effectively. Difference spectra revealed the presence of cytochromes (a + a3), b and c. Respiratory control was stimulated 2-fold by ADP with different exogenous oxidizable substrates. Calculated ADP/O ratios were consistent with the notion that ascorbate/N,N,N',N'-tetramethylphenylenediamine (TMPD)-linked and FAD-linked respiration proceeds, respectively, with one third and two thirds of the ATP producing capacity of NADH-linked respiration. State 3 was suppressed by the ATP synthase inhibitors oligomycin and aurovertin and by the adenine nucleotide translocator inhibitors atractyloside and carboxy atractyloside. The protonophore carbonyl cyanide p-(trifluoromethoxy)phenylhydrazone (FCCP) provoked state 3u respiration. The mitochondrial preparation was capable of Ca2+ uptake and Ca2+ stimulated respiration. Data obtained suggests strongly that mitochondrial complexes I, II, III and IV are present in a major pathway of electron-transfer and that oxidative phosphorylation might proceed with high bioenergetic efficiency.
Mol Biochem Parasitol 1997 Dec 01
PMID:Characterization of mitochondrial electron-transfer in Leishmania mexicana. 949 31

Fourteen Rhodobacter capsulatus mutants unable to grow with xanthine as sole nitrogen source were isolated by random Tn5 mutagenesis. Five of these Tn5 insertions were mapped within two adjacent chromosomal EcoRI fragments hybridizing to oligonucleotides synthesized according to conserved amino acid sequences of eukaryotic xanthine dehydrogenases. DNA sequence analysis of this region revealed two open reading frames, designated xdhA and xdhB, encoding xanthine dehydrogenase. The deduced amino acid sequence of XDHA contains binding sites for two [2Fe-2S] clusters and FAD, whereas XDHB is predicted to contain the molybdopterin cofactor. In contrast to R. capsulatus, these three cofactor binding sites reside within a single polypeptide chain in eukaryotic xanthine dehydrogenases. The amino acid sequence of xanthine dehydrogenase from R. capsulatus showed a higher degree of similarity to eukaryotic xanthine dehydrogenases than to the xanthine dehydrogenase-related aldehyde oxidoreductase from Desulphovibrio gigas. The expression of an xdhA-lacZ fusion was induced when hypoxanthine or xanthine was added as sole nitrogen source. Mutations in nifR1 (ntrC) and nifR4 (rpoN, encoding sigma54) had no influence on xdh gene expression. A putative activator sensing the availability of substrate seems to respond to xanthine but not to hypoxanthine. The transcriptional start site of xdhA was mapped by primer extension analysis. Comparison with known promoter elements revealed no significant homology. Xanthine dehydrogenase from R. capsulatus was purified to homogeneity. The enzyme consists of two subunits with molecular masses of 85 kDa and 50 kDa respectively. N-terminal amino acid sequencing of both subunits confirmed the predicted start codons. The molecular mass of the native enzyme was determined to be 275 kDa, indicating an alpha2beta2-subunit structure. Analysis of the molybdenum cofactor of xanthine dehydrogenase from R. capsulatus revealed that it contains the molybdopterin cofactor and not a molybdopterin dinucleotide derivative.
Mol Microbiol 1998 Feb
PMID:Xanthine dehydrogenase from the phototrophic purple bacterium Rhodobacter capsulatus is more similar to its eukaryotic counterparts than to prokaryotic molybdenum enzymes. 951 10

An interesting flavoprotein-type monoamine oxidase (MAO) was recently isolated from Aspergillus niger and cloned [Schilling, B. & Lerch, K. (1995a) Biochim. Biophys. Acta 1243, 529-537; Schilling, B. & Lerch, K. (1995b) Mol. Gen. Genet. 247, 430-438]. The properties of this MAO, as well as a substantial part of its amino acid sequence, resemble those of both MAO A and B from higher animals, raising the possibility that it may be an evolutionary precursor of these mitochondrial enzymes. It differs from MAO A and B in several respects, however, including the fact that it is soluble and of peroxisomal location and that the FAD is non-covalently attached. We have overexpressed the fungal enzyme (MAO-N) in Escherichia coli and isolated it in pure form. Since several of the observations of previous workers on MAO-N could not be reproduced, we have reexamined its substrate specificity, interaction with reversible and irreversible inhibitors and other catalytic and molecular properties. MAO-N has a considerably higher turnover number on many aliphatic and aromatic amines than either form of the mammalian enzyme. Some aspects of the substrate specificity resemble those of MAO B, while others are similar to MAO A, including biphasic kinetics in double reciprocal plots. Contrary to a previous report [Schilling, B. & Lerch, K. (1995a) Biochim. Biophys. Acta 1243, 529-537], however, the fungal enzyme does not oxidize serotonin, norepinephrine, dopamine or other biogenic amines. MAO-N is irreversibly inhibited by stoichiometric amounts of both (-)deprenyl and clorgyline in a mechanism-based reaction, forming flavocyanine adducts with N5 of the FAD, like the mammalian enzymes, but inactivation is much faster with clorgyline than deprenyl, suggesting a closer resemblance to MAO A than B. The dissociation constants for a large number of reversible competitive inhibitors have been determined for MAO-N and comparison with similar values for MAO A and B again pointed to a greater similarity to the former than the latter.
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PMID:Isolation and characterization of an evolutionary precursor of human monoamine oxidases A and B. 957 86

Monoamine oxidase B (MAO B) catalyzes the oxidative deamination of biogenic and xenobiotic amines. The oxidative step is coupled to the reduction of an obligatory cofactor, FAD, which is covalently linked to the apoenzyme at Cys397. Our previous studies identified two noncovalent flavin-binding regions in MAO B (residues 6-34 and 39-46) (Kwan, S.-W., Lewis, D. A., Zhou, B. P., and Abell, C. W. (1995) Arch. Biochem. Biophys. 316, 385-391; Zhou, B. P., Lewis, D. A., Kwan, S.-W., Kirksey, T. J., and Abell, C. W. (1995) Biochemistry 34, 9526-9531). In these regions, Glu34 and Tyr44 were found to be required for the initial binding of FAD. By comparing sequences with enzymes in the oxidoreductase family, we now have found an additional FAD-binding site in MAO B (residues 222-227), which is highly conserved across species (human, bovine, and rat). This conserved sequence contains adjacent glycine and aspartate residues (Gly226 and Asp227). Based on the x-ray crystal structures of several oxidoreductases (Eggink, G., Engel, H., Vriend, G., Terpstra, P., and Witholt, B. (1990) J. Mol. Biol. 212, 135-142; Van Driessche, G., Kol, M., Chen, Z.-W., Mathews, F. S., Meyer, T. E., Bartsch, R. G., Cusanovich, M. A., and Van Beeumen, J. J. (1996) Protein Sci. 5, 1753-1764), the Gly residue at the end of a beta-strand facilitates a sharp turn and extends the beta-carbonyl group of Asp to interact with the 3'-hydroxyl group of the ribityl chain of FAD. To assess the hypothesis that Gly226 and Asp227 are involved in FAD binding in MAO B, site-specific mutants that encode substitutions at these positions were prepared and expressed in mammalian COS-7 cells. Our results indicate that Gly226 and the beta-carbonyl group of Asp227 are required for covalent flavinylation and catalytic activity of MAO B, but not for noncovalent binding of FAD. Our studies also reveal that mutagenesis at Glu34 and Tyr44 not only interferes with covalent flavinylation and catalytic activity of MAO B, but also with noncovalent binding of FAD. Based on these collective results, we propose that the coupling of FAD to the MAO B apoenzyme is a multistep process.
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PMID:Characterization of a highly conserved FAD-binding site in human monoamine oxidase B. 961 88

The present study provides evidence that the reductive dechlorination of DDT (p, p'-dichlorodiphenyltrichloroethane) to DDD (p, p'-dichlorodiphenyldichloroethane) mediated by rat blood proceeds in the presence of both a reduced pyridine nucleotide and a flavin. The reduction appears to proceed in two steps. The first step is reduction of a flavin such as FAD, FMN or riboflavin by NADPH or NADH, either enzymatically or nonenzymatically. The second step is nonenzymatic reductive dechlorination of DDT by the reduced flavin, catalyzed by the heme group of hemoglobin.
Biochem Mol Biol Int 1998 Jun
PMID:Reductive dechlorination of DDT to DDD by rat blood. 963 32

The complex structure of glucose oxidase (GOX) with the substrate glucose was determined using a docking algorithm and subsequent molecular dynamics simulations. Semiempirical quantum chemical calculations were used to investigate the role of the enzyme and FAD co-enzyme in the catalytic oxidation of glucose. On the basis of a small active site model, substrate binding residues were determined and heats of formation were computed for the enzyme substrate complex and different potential products of the reductive half reaction. The influence of the protein environment on the active site model was estimated with a point charge model using a mixed QM/MM method. Solvent effects were estimated with a continuum model. Possible modes of action are presented in relation to experimental data and discussed with respect to related enzymes. The calculations indicate that the redox reaction of GOX differs from the corresponding reaction of free flavins as a consequence of the protein environment. One of the active site histidines is involved in substrate binding and stabilization of potential intermediates, whereas the second histidine is a proton acceptor. The former one, being conserved in a series of oxidoreductases, is also involved in the stabilization of a C4a-hydroperoxy dihydroflavin in the course of the oxidative half reaction.
J Comput Aided Mol Des 1998 Sep
PMID:Aspects of the mechanism of catalysis of glucose oxidase: a docking, molecular mechanics and quantum chemical study. 983 5


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