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Flavin Adenine Dinucleotide (flavin + adenine_dinucleotide)

Distribution by Scientific Domains

Chemistry	50%
Life Sciences	37%
Medical Sciences	12%

Selected Abstracts

Interaction of Flavin Adenine Dinucleotide (FAD) with a Glassy Carbon Electrode Surface

CHEMISTRY & BIODIVERSITY, Issue 8 2008
Haizhen Wei
Abstract The interaction of flavin adenine dinucleotide (FAD) with a glassy carbon electrode (GCE) surface was investigated in terms of the FAD adsorption thermodynamics and kinetics, the subsequent electroreduction mechanism, and the corresponding electron-transfer rate. The kinetics of FAD electroreduction at the GCE was found to be an adsorption-controlled process. A set of electroreduction kinetic parameters was calculated: the true number of electrons involved in the FAD reduction, n=1.76, the apparent transfer coefficient, ,app=0.41, and the apparent heterogeneous electron-transfer rate constant, kapp=1.4,s,1. The deviation of the number of exchanged electrons from the theoretical value for the complete reduction of FAD to FADH2 (n=2) indicates that a small portion of FAD goes to a semiquinone state during the redox process. The FAD adsorption was well described by the Langmuir adsorption isotherm. The large negative apparent Gibbs energy of adsorption (,Gads=,39.7 ±0.4,kJmol,1) indicated a highly spontaneous and strong adsorption of FAD on the GCE. The energetics of the adsorption process was found to be independent of the electrode surface charge in the electrochemical double-layer region. The kinetics of FAD adsorption was modeled using a pseudo -first-order kinetic model. [source]

Compound heterozygosity of two missense mutations in the NADH-cytochrome b5 reductase gene of a Polish patient with type I recessive congenital methaemoglobinaemia

EUROPEAN JOURNAL OF HAEMATOLOGY, Issue 6 2003
Dorota Grabowska
Abstract: A case of type I methaemoglobinaemia observed in a Polish subject with compound heterozygosity for two mutations in the reduced nicotinamide adenine dinucleotide (NADH) cytochrome b5 reductase (b5R) gene is described. One is a novel mutation 647T,C which leads to substitution of isoleucine by threonine at position 215 (I215T). This maternal mutation was found in several family members. A previously known mutation, 757G,A, leads to the replacement of valine by methionine at position 252 (V252M). The latter mutation was found also in the father and one of the two brothers. The effects of these mutations were analysed on a model of the human b5R protein obtained by homology modelling. Although both amino acid substitutions are located in the NADH-binding domain, the whole protein structure, especially the region between the flavin adenine dinucleotide and NADH-binding domains, is disturbed. The structural changes in the I215T mutant are less prominent than those in the V252M mutant. We presume that the 647T,C mutation is a type I mutation, however, it has not been observed in the homozygous state. [source]

Photoinduced electron transfer in glucose oxidase: a picosecond time-resolved ultraviolet resonance Raman study

JOURNAL OF RAMAN SPECTROSCOPY, Issue 11 2008
Akiko Fujiwara
Abstract Picosecond time-resolved ultraviolet resonance Raman (UVRR) spectroscopy has been applied to photoinduced electron transfer (ET) of glucose oxidase (GOD). In this study, we succeeded in directly observing changes in the aromatic amino acid residues in the photoinduced ET of GOD for the first time. UVRR spectra excited at 226 nm showed bands from Trp and Tyr residues. An intensity decrease of the Trp UVRR bands and the appearance of the UVRR bands attributable to Trp,+ were observed in the time-resolved spectra. In the time-resolved UVRR spectra excited at 240 nm, the intensity decrease of the flavin adenine dinucleotide (FAD) bands was also observed on the same time scale. These results showed that the Trp residue(s) serves as an electron donor to excited-state FAD in the photoinduced ET of GOD. The comparison of the temporal changes of the Trp and FAD band intensities suggested that the ET from the Trp residue(s) to the FAD occurs with a time constant of ,1.5 ps. Copyright © 2008 John Wiley & Sons, Ltd. [source]

Resonance Raman spectra of the neutral and anionic radical semiquinones of flavin adenine dinucleotide in glucose oxidase revisited

JOURNAL OF RAMAN SPECTROSCOPY, Issue 8 2006
Johannes P. M. Schelvis
Abstract Flavin radical semiquinones are intermediates in important physiological processes. Resonance Raman (RR) spectroscopy is an important tool to determine the interactions between these radical intermediates and their protein environment that regulate their reactivity and role in the reaction mechanisms. RR spectra of flavin radical semiquinones have been available for several flavoproteins, and those in the glucose oxidase (GO) seem significantly different from all the other available data. Since GO is often used not only as a standard for flavin-containing proteins but also in biotechnological applications, we decided to reexamine the RR spectra of the neutral and anionic radical semiquinone forms of the flavin adenine dinucleotide (FAD) cofactor in this enzyme. The new data show that the vibrational wavenumbers of the neutral and anionic radical semiquinone forms of FAD in GO are very similar to those in other flavoproteins. The discrepancies that were observed earlier seem related to contributions of the FAD in different redox and protonation states. We also obtained the first RR spectra of the oxidized FAD cofactor in GO. Analysis of the vibrations of the oxidized FAD and its anionic radical semiquinone in GO in H2O and D2O solutions indicates that the subtle differences between these spectra in GO and in other flavoproteins are related to the weak hydrogen-bonding environment of the FAD cofactor in GO. Copyright © 2006 John Wiley & Sons, Ltd. [source]

Flavin-sensitized Photo-oxidation of Lysozyme and Serum Albumin

PHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 4 2009
Yazhou Zhang
The excited state processes of riboflavin, flavin mononucleotide and flavin adenine dinucleotide in argon-saturated aqueous solution were studied in the presence of lysozyme or bovine serum albumin (BSA). UV,Vis absorption and fluorescence spectroscopy indicates that the noncovalent flavin-protein binding is relatively weak. Quenching of the flavin triplet state by BSA, observed by time-resolved photolysis, is less efficient than by lysozyme. Light-induced oxidation of the two proteins and reduction of the three flavins were studied. The quantum yields of the former and latter in the absence of oxygen are up to 0.1 and 0.04, respectively. The effects of pH and sensitizer and protein concentrations were examined in greater detail. The proposed reaction is electron transfer from the tryptophan moiety to the flavin triplet rather than excited singlet state. [source]

Involvement of Electron Transfer in the Photoreaction of Zebrafish Cryptochrome-DASH,

PHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 4 2008
Kazunori Zikihara
Photoreaction of a blue-light photoreceptor Cryptochrome-DASH (Cry-DASH), a new member of the Cryptochrome family, from zebrafish was studied by UV,visible absorption spectroscopy in aqueous solutions at 293 K. Zebrafish Cry-DASH binds two chromophores, a flavin adenine dinucleotide (FAD) and a N5,N10-methenyl-5,6,7,8-tetrahydrofolate (MTHF) noncovalently. The bound FAD exists in the oxidized form (FADox) in the dark. Blue light converts FADox to the neutral radical form (FADH?). Formed FADH? is transformed to the fully reduced form FADH2 (or FADH,) by successive light irradiation, or reverts to FADox. FADH2 (or FADH,) reverts to FADH? or possibly to FADox directly. The effect of dithiothreitol suggests a possible electron transfer between FAD in zebrafish Cry-DASH and reductants in the external medium. This is the first report on the photoreaction pathway and kinetics of a vertebrate Cry-DASH family protein. [source]

Endogenous Fluorescence Spectroscopy of Cell Suspensions for Chemopreventive Drug Monitoring,

PHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 1 2005
Nathaniel D. Kirkpatrick
ABSTRACT Cancer chemopreventive agents such as N -4-(hydroxyphenyl)-retinamide (4HPR) are thought to prevent cancers by suppressing growth or inducing apoptosis in precancerous cells. Mechanisms by which these drugs affect cells are often not known, and the means to monitor their effects is not available. In this study endogenous fluorescence spectroscopy was used to measure metabolic changes in response to treatment with 4HPR in ovarian and bladder cancer cell lines. Fluorescence signals consistent with nicotinamide adenine dinucleotide (NADH), flavin adenine dinucleotide (FAD) and tryptophan were measured to monitor cellular activity through redox status and protein content. Cells were treated with varying concentrations of 4HPR and measured in a stable environment with a sensitive fluorescence spectrometer. Results suggest that redox signal of all cells changed in a similar dose-dependant manner but started at different baseline levels. Redox signal changes depended primarily on changes consistent with NADH fluorescence, whereas the FAD fluorescence remained relatively constant. Similarly, tryptophan fluorescence decreased with increased drug treatment, suggesting a decrease in protein production. Given that each cell line has been shown to have a different apoptotic response to 4HPR, fluorescence redox values along with changes in tryptophan fluorescence may be a response as well as an endpoint marker for chemopreventive drugs. [source]

Characterization of two members of the cryptochrome/photolyase family from Ostreococcus tauri provides insights into the origin and evolution of cryptochromes

PLANT CELL & ENVIRONMENT, Issue 10 2010
MARC HEIJDE
ABSTRACT Cryptochromes (Crys) are blue light receptors believed to have evolved from the DNA photolyase protein family, implying that light control and light protection share a common ancient origin. In this paper, we report the identification of five genes of the Cry/photolyase family (CPF) in two green algae of the Ostreococcus genus. Phylogenetic analyses were used to confidently assign three of these sequences to cyclobutane pyrimidine dimer (CPD) photolyases, one of them to a DASH-type Cry, and a third CPF gene has high homology with the recently described diatom CPF1 that displays a bifunctional activity. Both purified OtCPF1 and OtCPF2 proteins show non-covalent binding to flavin adenine dinucleotide (FAD), and additionally to 5,10-methenyl-tetrahydrofolate (MTHF) for OtCPF2. Expression analyses revealed that all five CPF members of Ostreococcus tauri are regulated by light. Furthermore, we show that OtCPF1 and OtCPF2 display photolyase activity and that OtCPF1 is able to interact with the CLOCK:BMAL heterodimer, transcription factors regulating circadian clock function in other organisms. Finally, we provide evidence for the involvement of OtCPF1 in the maintenance of the Ostreococcus circadian clock. This work improves our understanding of the evolutionary transition between photolyases and Crys. [source]

Temperature dependence of nitrate reductase in the psychrophilic unicellular alga Koliella antarctica and the mesophilic alga Chlorella sorokiniana

PLANT CELL & ENVIRONMENT, Issue 7 2006
VITTORIA DI MARTINO RIGANO
ABSTRACT Temperature responses of nitrate reductase (NR) were studied in the psychrophilic unicellular alga, Koliella antarctica, and in the mesophilic species, Chlorella sorokiniana. Enzymes from both species were purified to near homogeneity by Blue Sepharose (Pharmacia, Uppsala, Sweden) affinity chromatography and high-resolution anion-exchange chromatography (MonoQ; Pharmacia; Uppsala, Sweden). Both enzymes have a subunit molecular mass of 100 kDa, and K. antarctica NR has a native molecular mass of 367 kDa. NR from K. antarctica used both NADPH and NADH, whereas NR from C. sorokiniana used NADH only. Both NRs used reduced methyl viologen (MVH) or benzyl viologen (BVH). In crude extracts, maximal NADH and MVH-dependent activities of cryophilic NR were found at 15 and 35 °C, respectively, and retained 77 and 62% of maximal activity, respectively, at 10 °C. Maximal NADH and MVH-dependent activities of mesophilic NR, however, were found at 25 and 45 °C, respectively, with only 33 and 23% of maximal activities being retained at 10 °C. In presence of 2 µm flavin adenine dinucleotide (FAD), activities of cryophilic NADH:NR and mesophilic NADH:NR were stable up to 25 and 35 °C, respectively. Arrhenius plots constructed with cryophilic and mesophilic MVH:NR rate constants, in both presence or absence of FAD, showed break points at 15 and 25 °C, respectively. Essentially, similar results were obtained for purified enzymes and for activities measured in crude extracts. Factors by which the rate increases by raising temperature 10 °C (Q10) and apparent activation energy (Ea) values for NADH and MVH activities measured in enzyme preparations without added FAD differed slightly from those measured with FAD. Overall thermal features of the NADH and MVH activities of the cryophilic NR, including optimal temperatures, heat inactivation (with/without added FAD) and break-point temperature in Arrhenius plots, are all shifted by about 10 °C towards lower temperatures than those of the mesophilic enzyme. Transfer of electrons from NADH to nitrate occurs via all three redox centres within NR molecule, whereas transfer from MVH requires Mo-pterin prosthetic group only; therefore, our results strongly suggest that structural modification(s) for cold adaptation affect thermodynamic properties of each of the functional domains within NR holoenzyme in equal measure. [source]

Sequence-structure analysis of FAD-containing proteins

PROTEIN SCIENCE, Issue 9 2001
Orly Dym
We have analyzed structure-sequence relationships in 32 families of flavin adenine dinucleotide (FAD)-binding proteins, to prepare for genomic-scale analyses of this family. Four different FAD-family folds were identified, each containing at least two or more protein families. Three of these families, exemplified by glutathione reductase (GR), ferredoxin reductase (FR), and p -cresol methylhydroxylase (PCMH) were previously defined, and a family represented by pyruvate oxidase (PO) is newly defined. For each of the families, several conserved sequence motifs have been characterized. Several newly recognized sequence motifs are reported here for the PO, GR, and PCMH families. Each FAD fold can be uniquely identified by the presence of distinctive conserved sequence motifs. We also analyzed cofactor properties, some of which are conserved within a family fold while others display variability. Among the conserved properties is cofactor directionality: in some FAD-structural families, the adenine ring of the FAD points toward the FAD-binding domain, whereas in others the isoalloxazine ring points toward this domain. In contrast, the FAD conformation and orientation are conserved in some families while in others it displays some variability. Nevertheless, there are clear correlations among the FAD-family fold, the shape of the pocket, and the FAD conformation. Our general findings are as follows: (a) no single protein ,pharmacophore' exists for binding FAD; (b) in every FAD-binding family, the pyrophosphate moiety binds to the most strongly conserved sequence motif, suggesting that pyrophosphate binding is a significant component of molecular recognition; and (c) sequence motifs can identify proteins that bind phosphate-containing ligands. [source]

Crystallization and preliminary crystallographic studies of a flavin-dependent thymidylate synthase from Helicobacter pylori

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 5 2010
Xiaoli Zhang
The ThyX enzymes that have recently been identified in various bacteria, including some important human pathogens such as Helicobacter pylori and Mycobacterium tuberculosis, are flavin-dependent thymidylate synthases that function in the place of classic thymidylate synthase enzymes in the biosynthesis of dTMP, which is one of the building blocks of DNA. They are promising targets for the development of novel antibiotics because they utilize catalytic mechanisms that are distinct from those of the classic thymidylate synthases found in most organisms, including humans. In this study, H. pylori ThyX was purified and crystallized in complex with flavin adenine dinucleotide (FAD) and a diffraction data set was collected to 2.5,Å resolution. The crystals belonged to space group C2, with unit-cell parameters a = 221.92, b = 49.43, c = 143.02,Å, , = 98.84°. [source]

The structures of pyruvate oxidase from Aerococcus viridans with cofactors and with a reaction intermediate reveal the flexibility of the active-site tunnel for catalysis

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 11 2007
Ella Czarina Magat Juan
The crystal structures of pyruvate oxidase from Aerococcus viridans (AvPOX) complexed with flavin adenine dinucleotide (FAD), with FAD and thiamine diphosphate (ThDP) and with FAD and the 2-acetyl-ThDP intermediate (AcThDP) have been determined at 1.6, 1.8 and 1.9,Å resolution, respectively. Each subunit of the homotetrameric AvPOX enzyme consists of three domains, as observed in other ThDP-dependent enzymes. FAD is bound within one subunit in the elongated conformation and with the flavin moiety being planar in the oxidized form, while ThDP is bound in a conserved V-conformation at the subunit,subunit interface. The structures reveal flexible regions in the active-site tunnel which may undergo conformational changes to allow the entrance of the substrates and the exit of the reaction products. Of particular interest is the role of Lys478, the side chain of which may be bent or extended depending on the stage of catalysis. The structures also provide insight into the routes for electron transfer to FAD and the involvement of active-site residues in the catalysis of pyruvate to its products. [source]

Recessive congenital methaemoglobinaemia: functional characterization of the novel D239G mutation in the NADH-binding lobe of cytochrome b5 reductase

BRITISH JOURNAL OF HAEMATOLOGY, Issue 6 2005
M. J. Percy
Summary Type I recessive congenital methaemoglobinaemia (RCM), caused by the reduced form of nicotinamide adenine dinucleotide (NADH)-cytochrome b5 reductase (cytb5r) deficiency, manifests clinically as cyanosis without neurological dysfunction. Two mutations, E255- and G291D, have been identified in the NADH-binding lobe of cytb5r in previously reported patients, and we have detected a further novel mutation, D239G, in this lobe in two unrelated Irish families. Although one family belongs to the genetically isolated Traveller Community, which separated from the general Irish population during the 1845,48 famine, the D239G mutation was present on the same haplotype in both families. Three known cytb5r mutations were also identified, including the R159- mutation, which causes loss of the entire NADH-binding lobe and had previously been reported in an individual with type II RCM. Characterization of the three NADH-binding lobe mutants using a heterologous expression system revealed that all three variants retained stoichiometric levels of flavin adenine dinucleotide with spectroscopic and thermodynamic properties comparable with those of native cytb5r. In contrast to the E255- and G291D variants, the novel D239G mutation had no adverse impact on protein thermostability. The D239G mutation perturbed substrate binding, causing both decreased specificity for NADH and increased specificity for NADPH. Thus cytb5r deficient patients who are heterozygous for an NADH-binding lobe mutation can exhibit the clinically less severe type I phenotype, even in association with heterozygous deletion of the NADH-binding lobe. [source]

Insight into the Metabolism Rate of Quinone Analogues from Molecular Dynamics Simulation and 3D-QSMR Methods

CHEMICAL BIOLOGY & DRUG DESIGN, Issue 4 2007
Hai-Feng Chen
Molecular dynamics simulation was applied to investigate the metabolism mechanism for quinone analogues. Favourable hydrogen bonds between ligand and NQO1, and parallel orientation between ligand and flavin adenine dinucleotide could explain the difference of metabolism rate (in ,mol/min/mg) for quinone analogues. This is consistent with the experimental observation (Structure 2001;9:659,667). Then Support Vector Machines was used to construct quantitative structure,metabolism rate model. The model was evaluated by 14 test set compounds. Some descriptors selected by Support Vector Machine, were introduced into standard fields of three-dimensional quantitative structure,metabolism relationship to improve the statistical parameters of three-dimensional quantitative structure,metabolism relationship models. The results show that the inclusion of highest occupied molecular orbital and lowest unoccupied molecular orbital is meaningful for three-dimensional quantitative structure,metabolism relationship models. These in silico absorption, distribution, metabolism and excretion models are helpful in making quantitative prediction of their metabolic rates for new lead compounds before resorting in vitro and in vivo experimentation. [source]