共查询到20条相似文献,搜索用时 15 毫秒
1.
Copper amine oxidases catalyze the oxidative deamination of primary amines operating through a ping-pong bi bi mechanism, divided into reductive and oxidative half-reactions. Considerable debate still exists regarding the role of copper in the oxidative half-reaction, where O2 is reduced to H2O2. Substrate-reduced amine oxidases display an equilibrium between a Cu(II) aminoquinol and a Cu(I) semiquinone, with the magnitude of the equilibrium constant being dependent upon the enzyme source. The initial electron transfer to dioxygen has been proposed to occur from either the reduced Cu(I) center or the reduced aminoquinol cofactor. In order for Cu(I) to be involved, it must be shown that the rate of electron transfer (k
ET) between the aminoquinol and Cu(II) is sufficiently rapid to place the Cu(I) semiquinone moiety on the mechanistic pathway. To further explore this issue, we measured the intramolecular electron transfer rate for the Cu(II) aminoquinol ⇆ Cu(I) semiquinone equilibrium in Arthrobacter globiformis amine oxidase (AGAO) by temperature-jump relaxation techniques. The results presented herein establish that k
ET is greater than the rate of catalysis (k
cat) for the preferred amine substrate β-phenylethylamine at three pH values, thereby permitting the Cu(I) semiquinone to be a viable catalytic intermediate during enzymatic reoxidation in this enzyme. The data show that k
ET is approximately equivalent at pH 6.2 and 7.2, being 2.5 times k
cat for these pH values. At pH 8.2, however, k
ET decreases, becoming comparable to k
cat. Potential reasons for the decreased k
ET at basic pH are presented. The implications of these results in light of a previously published study measuring reoxidation rates of substrate-reduced AGAO are also addressed. 相似文献
2.
Mark P. Reynolds Andrew J. Baron Carrie M. Wilmot Elinor Vinecombe Conrad Stevens Simon E. V. Phillips Peter F. Knowles M. J. McPherson 《Journal of biological inorganic chemistry》1997,2(3):327-335
The catalytic mechanism of the copper-containing enzyme galactose oxidase involves a protein radical on Tyr272, one of the
equatorial copper ligands. The first step in this mechanism has been proposed to be the abstraction of a proton from the alcohol
substrate by Tyr495, the axial copper ligand that is weakly co-ordinated to copper. In this study we have generated and studied
the properties of a Y495F variant to test this proposal. X-ray crystallography reveals essentially no change from wild-type
other than loss of the tyrosyl hydroxyl group. Visible spectroscopy indicates a significant change in the oxidised Y495F compared
to wild-type with loss of a broad 810-nm peak, supporting the suggestion that this feature is due to inter-ligand charge transfer
via the copper. The presence of a peak at 420 nm indicates that the Y495F variant remains capable of radical formation, a
fact supported by EPR measurements. Thus the significantly reduced catalytic efficiency (1100-fold lower k
cat / K
m) observed for this variant is not due to an inability to generate the Tyr272 radical. By studying azide-induced pH changes,
it is clear that the reduced catalytic efficiency is due mainly to the inability of Y495F to accept protons. This provides
definitive evidence for the key role of Tyr495 in the initial proton abstraction step of the galactose oxidase catalytic mechanism.
Received: 17 December 1996 / Accepted: 12 March 1997 相似文献
3.
The interactions of cyanide with two copper-containing amine oxidases (CuAOs) from pea seedlings (PSAO) and the soil bacterium Arthrobacter globiformis (AGAO) have been investigated by spectroscopic and kinetic techniques. Previously, we rationalized the effects of azide and cyanide for several CuAOs in terms of copper coordination by these exogenous ligands and their effects on the internal redox equilibrium TPQamr-Cu(II)TPQsq-Cu(I). The mechanism of cyanide inhibition was proposed to occur through complexation to Cu(I), thereby directly competing with O2 for reoxidation of TPQ. Although cyanide readily and reversibly reacts with quinones, no direct spectroscopic evidence for cyanohydrin derivatization of TPQ has been previously documented for CuAOs. This work describes the first direct spectroscopic evidence, using both model and enzyme systems, for cyanohydrin derivatization of TPQ. Kd values for Cu(II)-CN– and Cu(I)-CN–, as well as the Ki for cyanide inhibition versus substrate amine, are reported for PSAO and AGAO. In spite of cyanohydrin derivatization of the TPQ cofactor in these enzymes, the uncompetitive inhibition of amine oxidation is determined to arise almost exclusively through CN– complexation of Cu(I).Abbreviations AGAO Arthrobacter globiformis amine oxidase - APAO Arthrobacter P1 amine oxidase - APT attached proton test - BPAO bovine plasma amine oxidase - CuAO quinone-copper containing amine oxidase - LTQ lysyl tyrosylquinone - MAO monoamine oxidase - PKAO porcine kidney amine oxidase - PPAO porcine plasma amine oxidase - PSAO pea seedling amine oxidase - TPQ 2,4,5-trihydroxyphenylalaninequinone - TPQamr TPQ aminoresorcinol - TPQimq TPQ iminoquinone - TPQox TPQ oxidized - TPQsq TPQ semiquinone - WT wild-typeE.M. Shepard and G.A. Juda contributed equally to this workThis revised version was published online in February 2004: Hansenula polymorpha was not italicised at the end of the Introduction, Equation 3 appeared twice, and the resolution of Scheme 3 was insufficient.An erratum to this article can be found at 相似文献
4.
Michele A. McGuirl Doreen E. Brown D. M. Dooley 《Journal of biological inorganic chemistry》1997,2(3):336-342
The interactions of five copper-containing amine oxidases with substrates and substrate analogues in the presence of the
copper ligands cyanide, azide, chloride, and 1,10-phenanthroline have been investigated. While cyanide inhibits, to varying
degrees, the reaction of phenylhydrazine with porcine kidney amine oxidase (PKAO), porcine plasma amine oxidase (PPAO), bovine
plasma amine oxidase (BPAO), and pea seedling amine oxidase (PSAO), it enhances the reaction of Arthrobacter P1 amine oxidase (APAO) with this substrate analogue. This indicates that cyanide exerts an indirect effect on topa quinone
(TPQ) reactivity via coordination to Cu(II) rather than through cyanohydrin formation at the TPQ organic cofactor. Moreover,
cyanide binding to the mechanistically relevant TPQ• semiquinone form of substrate-reduced APAO and PSAO was not observable by EPR or resonance Raman spectroscopy. Hence, cyanide
most likely inhibits enzyme reoxidation by binding to Cu(I) and trapping the Cu(I)-TPQ• form of amine oxidases, and thus preventing the reaction of O2 with Cu(I). In contrast, ligands such as azide, chloride, and 1,10-phenanthroline, which preferentially bind to Cu(II), inhibit
by stabilizing the aminoquinol Cu(II)-TPQred redox state, which is in equilibrium with Cu(I)-TPQ•.
Received: 12 December 1996 / Accepted: 20 March 1997 相似文献
5.
The intramolecular electron-transfer rate constant for the Cu(II)–topaNH2⇌ Cu(I)–topaSQ equilibrium in methylamine oxidase has been measured by temperature-jump relaxation techniques. At pH 7.0 the estimated kobs = 150±30 s–1 for both methylamine and benzylamine; assuming the equilibrium constant is ≈0.7–1 at pH 7.0 and 296 K, this would correspond
to a forward electron-transfer rate constant kET≈ 60–75 s–1. Although substantially slower than the previously determined kET≈ 20 000 s–1 for pea seedling amine oxidase [5] steady-state kinetics measurements established that kET > kcat≈ 4–10 s–1. Thus the Cu(I)-semiquinone state is a viable intermediate in methylamine oxidase turnover.
Received: 16 August 1995 / Accepted: 21 December 1995 相似文献
6.
Plant copper/quinone amine oxidases are homodimeric enzymes containing Cu(II) and a quinone derivative of a tyrosyl residue (2,4,5-trihydroxyphenylalanine, TPQ) as cofactors. These enzymes catalyze the oxidative deamination of primary amines by a classical ping-pong mechanism, i.e. two distinct half-reactions, enzyme reduction by substrate followed by its re-oxidation by molecular oxygen. In the first half-reaction two forms of the reduced TPQ have been observed, the colorless Cu(II)-aminoquinol and the yellow Cu(I)-semiquinolamine radical so that this enzyme may be referred to as a "protein-radical enzyme". The interaction of xenon, in aqueous solutions, with the copper/TPQ amine oxidase from lentil (Lens esculenta) seedlings has been investigated by NMR and optical spectroscopy. NMR data indicate that xenon binds to the protein. Under 10 atm gaseous xenon and in the absence of substrates more than 60% native enzyme is converted into Cu(I)-semiquinolamine radical species, showing for the first time that both monomers in the dimer can generate the radical. Under the same experimental conditions the copper-free lentil enzyme is able to generate an intermediate absorbing at about 360 nm, which is assigned to the product Schiff base quinolaldimine which, to the best of our knowledge, has never been observed during the catalytic mechanism of plant amine oxidases. A possible role of the lysine residue responsible for the formation of Cu(I)-semiquinolamine and quinolaldimine, is proposed. 相似文献
7.
Abstract Arthrobacter globiformis amine oxidase produced by Escherichia coli cells grown in copper-depleted media was reported to undergo activation due to formation of its topaquinone cofactor in a copper-dependent autocatalytic reaction. Likewise, a mutated E. coli amine oxidase located in the cytoplasm was reported to form topaquinone autocatalytically in an EDTA-sensitive reaction. Here we show unequivocally that formation of an amine oxidase lacking topaquinone is primarily a consequence of the location of the enzyme in the cytoplasm rather than the level of copper in the growth medium. For E. coli , insertion of copper into apoamine oxidase and subsequent topaquinone formation occur after export of the apoenzyme into the periplasm. 相似文献
8.
9.
LodA is a novel lysine-ε-oxidase which possesses a cysteine tryptophylquinone cofactor. It is the first tryptophylquinone enzyme known to function as an oxidase. A steady-state kinetic analysis shows that LodA obeys a ping-pong kinetic mechanism with values of kcat of 0.22 ± 0.04 s−1, Klysine of 3.2 ± 0.5 μM and KO2 of 37.2 ± 6.1 μM. The kcat exhibited a pH optimum at 7.5 while kcat/Klysine peaked at 7.0 and remained constant to pH 8.5. Alternative electron acceptors could not effectively substitute for O2 in the reaction. A mechanism for the reductive half reaction of LodA is proposed that is consistent with the ping-pong kinetics. 相似文献
10.
Highly polymorphic microsatellites of rice consist of AT repeats, and a classification of closely related cultivars with these microsatellite loci 总被引:20,自引:3,他引:20
H. Akagi Y. Yokozeki A. Inagaki T. Fujimura 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1997,94(1):61-67
Microsatellites consisting of AT repeats are highly polymorphic in rice genomes and can be used to distinguish between even
closely related japonica cultivars in Japan. Polymorphisms of 20 microsatellite loci were determined using 59 japonica cultivars,
including both domestic and modern Japanese cultivars. Although the polymorphisms of these 20 microsatellite loci indicated
that the Japanese cultivars were genetically quite similar, microsatellites consisting of AT repeats showed high gene diversity
even among such closely related cultivars. Combinations of these hypervariable microsatellites can be employed to classify
individual cultivars, since the microsatellites were stable within each cultivar. An identification system based on these
highly polymorphic microsatellites could be used to maintain the purity of rice seeds by eliminating contamination. A parentage
diagnosis using 17 polymorphic microsatellite loci clearly demonstrated that plants which carried desired chromosome regions
had been selected in breeding programs. Thus, these hypervariable microsatellites consisting of AT repeats should promote
the selection of plants which carry desired chromosomes from genetically similar parents. Backcrossing could also help to
eliminate unnecessary chromosome regions with microsatellite polymorphisms at an early stage in breeding programs.
Received: 8 July 1996 / Accepted: 12 July 1996 相似文献
11.
Dacil Zurita Isabelle Gautier-Luneau Stéphane Ménage J.-L. Pierre Eric Saint-Aman 《Journal of biological inorganic chemistry》1997,2(1):46-55
Copper(II) complexes derived from the tripodal ligand bis(3′-t–butyl-2′-hydroxybenzyl)(2-pyridylmethyl)amine (LH2) have been studied in order to mimic the redox active site of the free radical-containing copper metalloenzyme galactose
oxidase. In non-coordinating solvents such as dichloromethane, only an EPR-silent dimeric complex was obtained (L2Cu2). The crystal structure of L2Cu2 revealed a "butterfly" design of the [Cu(μOR)2Cu] unit, which is not flattened and leads to a short Cu–Cu distance, the t–butyl groups being localized on the same side of the [Cu(μOR)2Cu] unit. The dimeric structure was broken down by acetonitrile or by alcohols, leading quantitatively to a brown mononuclear
copper(II) complex. UV-visible and EPR data indicated the coordination of the solvent in these mononuclear complexes. Electrochemical
as well as chemical (silver acetate) one-electron oxidation of acetonitrile solutions of the monomeric complex led to a yellow-green
solution. Based on EPR, UV-visible and resonance Raman spectroscopy, the one-electron oxidation product was identified as
a cupric phenoxyl radical system. It slowly decomposes into a product where the ligand has been substituted (dimerization)
in the para position of the hydroxyl group, for one of the phenolic groups. The data for the one-electron oxidized species provides strong
evidence for a free-radical copper (II) complex.
Received: 19 July 1996 / Accepted: 16 October 1996 相似文献
12.
In this paper, we report the inactivation of copper containing bovine plasma amine oxidase (BPAO) by a series of saturated alkylamines containing halogen atoms at γ-position, which are 1,1,1-trihalo-3-aminopropane, 1,1,1-trifluoro-2-hydroxy-3-aminopropane, 1,1,1-trichloro-2-hydroxy-3-aminopropane, and 1,1,1-trichloro-2-(2-phenethyloxy)-3-aminopropane. The trihalo-2-hydroxypropylamine analogs exhibited a time-dependent inactivation behavior of BPAO, with 1,1,1-trifluoro-2-hydroxy-3-aminopropane as the most efficient inactivator. The incorporation of a OH group at β-position increased inactivation efficiency by 10-fold within the trifluoro analogs, and the incorporation of a phenethyloxy group at β-position exhibited a higher efficiency by 3-fold within the trichloro analogs based on I75 values. All four compounds were found to be irreversible inactivators for BPAO. 相似文献
13.
Summary. In the paper here presented we summarize some results obtained in our laboratory in the last few years on new structural
and functional aspects of some amine oxidases (AOs), which have to be taken into consideration in defining new strategies
of controlling the cellular physiopathology.
In particular, the ability of Cu-AO purified from vegetal sources or from bovine serum to bind different cellular targets
inducing in them conformational as well as chemical modifications are described and the consequences of this interaction on
cellular functions are discussed. This is the case of the protective effect of Cu-AO against the damage induced by free radicals,
cell enrichment with Cu-AO, induction of cataract and the leukocyte-endothelia interaction.
The role of Cu and FAD-amine oxidases related as to the protection or damage of cells is also discussed. In this context the
involvement of MAOs in the modulation of the mitochondrial functions and in the induction of apoptosis is described and some
aspects of the molecular mechanism of AO inhibition by H2O2 and metronidazole analyzed.
Received January 9, 2002 Accepted June 27, 2002 Published online November 14, 2002
Acknowledgements This paper was supported by C.N.R. grant n° G002FD1 and MURST grant (Giovani Ricercatori, 2000).
Authors' address: Prof. Bruno Mondovì, Department of Biochemical Science, University of Rome “La Sapienza”, P.le Aldo Moro 5, I-00185 Rome,
Italy 相似文献
14.
Achim Sokolowski Heiko Leutbecher Thomas Weyhermüller Robert Schnepf Eberhard Bothe Eckhard Bill Peter Hildebrandt K. Wieghardt 《Journal of biological inorganic chemistry》1997,2(4):444-453
The reaction of the macrocycles 1,4,7-tris (3,5-di-tert-butyl-2-hydroxy-benzyl)-1,4,7-triazacyclononane, L1H3, or 1,4,7-tris(3-tert-butyl-5-methoxy-2-hydroxy-benzyl)-1,4,7-triazacyclononane, L2H3, with Cu(ClO4)2·6H2O in methanol (in the presence of Et3N) affords the green complexes [CuII(L1H)] (1), [CuII(L2H)]·CH3OH (2) and (in the presence of HClO4) [CuII(L1H2)](ClO4) (3) and [CuII(L2H2)] (ClO4) (4). The CuII ions in these complexes are five-coordinate (square-base pyramidal), and each contains a dangling, uncoordinated pendent
arm (phenol). Complexes 1 and 2 contain two equatorially coordinated phenolato ligands, whereas in 3 and 4 one of these is protonated, affording a coordinated phenol. Electrochemically, these complexes can be oxidized by one electron,
generating the phenoxyl-copper(II) species [CuII(L1H)]+ ·, [Cu(L2H)]+ ·, [CuII(L1H2)]2+ ·, and [CuII(L2H2)]2+ ·, all of which are EPR-silent. These species are excellent models for the active form of the enzyme galactose oxidase (GO).
Their spectroscopic features (UV-VIS, resonance Raman) are very similar to those reported for GO and unambiguously show that
the complexes are phenoxyl-copper(II) rather than phenolato-copper(III) species.
Received: 10 February 1997 / Accepted: 7 April 1997 相似文献
15.
《Journal of enzyme inhibition and medicinal chemistry》2013,28(5):311-325
AbstractIn this review, inhibitors of plant copper amine oxidases from Lens esculenta seedlings, Pisum sativum seedlings, and Euphorbia characias latex are described. Reversible competitive inhibitors and non-competitive inhibitors, irreversible active-site directed inhibitors and mechanism-based inactivators are reviewed in regard to their mechanisms of action. 相似文献
16.
The role of the polypeptide matrix in electron transfer processes in proteins has been studied in two distinct systems: first
in a protein where the induced ET is artificial, and second as part of the catalytic cycle of an enzyme. Azurins are structurally
well-characterized blue single-copper proteins consisting of a rigid β-sheet polypeptide matrix. We have determined rate constants
and activation parameters for intramolecular long-range electron transfer between the disulfide radical anions (generated
by pulse radiolysis) and the copper(II) centre as a function of driving force and nature of the intervening medium in a large
number of wild-type and single-site-mutated proteins. In ascorbate oxidase, for which the three-dimensional structure is equally
well characterized, the internal ET from the type-I Cu(I) to the trinuclear Cu(II) centre has been studied. We find that the
results correlate well with distance through well-defined pathways using a through-bond electron tunnelling mechanism.
Received: 2 January 1997 / Accepted: 6 February 1997 相似文献
17.
G. Musci Terri Z. L. Fraterrigo Lilia Calabrese David R. McMillin 《Journal of biological inorganic chemistry》1999,4(4):441-446
The possibility that ceruloplasmin (CP) functions as a copper transferase has fueled a continuing interest in studies of
the copper release process. The principal goal of the current investigation has been to identify the most labile copper centers
in sheep protein. In fact, subjecting the enzyme to a slow flux of cyanide at pH 5.2 under nitrogen in the presence of ascorbate
and a phenanthroline ligand produces partially demetalated forms of the protein. By standard chromatographic techniques it
is possible to isolate protein with a Cu/CP ratio of ∼4 or ∼5 as opposed to the native protein which has Cu/CP=5.8. In contrast
to other blue oxidases, analysis suggests that CP preferentially loses its type 1 coppers under these conditions. Thus, the
spectroscopic signals from the type 1 centers exhibit a loss of intensity while the EPR signal of the type 2 copper becomes
stronger. Furthermore, the Cu/CP≈4 and Cu/CP≈5 components retain about 50% of the activity of the native protein, consistent
with an intact type 2/type 3 cluster. All three type 1 copper sites appear to suffer copper loss. Reconstitution with a copper(I)
reagent restores the spectroscopic properties of the native protein and 90% of the original activity. The results suggest
a possible functional significance for the presence of three type 1 coppers in CP. By employing a pool of redox-active but
relatively labile type 1 copper centers, the enzyme can serve as a copper donor, if necessary, without completely sacrificing
its oxidase activity.
Received: 15 February 1999 / Accepted: 22 April 1999 相似文献
18.
19.
Mark D. Harrison Christopher E. Jones C. T. Dameron 《Journal of biological inorganic chemistry》1999,4(2):145-153
Copper is an absolute requirement for living systems and the intracellular trafficking of this metal to copper-dependent
proteins is fundamental to normal cellular metabolism. The copper chaperones perform the dual functions of trafficking and
the prevention of cytoplasmic exposure to copper ions in transit. Only a small number of copper chaperones have been identified
at this time but their conservation across plant, bacterial and animal species suggests that the majority of living systems
utilise these proteins for copper routing. The available data suggest that each copper-dependent protein in the cell is served
by a specific copper chaperone. Although copper chaperones cannot be substituted for one another in a given cell type, copper
chaperones that deliver to the same protein in different cell types appear to be functionally equivalent. The majority of
the copper chaperones identified thus far have an "open-faced β-sandwich" global fold with a conserved MXCXXC metal-binding
motif. Specificity for a given copper-dependent protein appears to be mediated by the residues surrounding the copper-binding
motif. Copper binds to such proteins as Cu(I) in a trigonal complex with three sulfur ligands. Only the copper chaperone specific
for cytochrome-c-oxidase, Cox17, deviates from this design.
Received: 12 October 1998 / Accepted: 7 December 1998 相似文献
20.
A. van Tuinen M. Koornneef M. -M. Cordonnier-Pratt L. H. Pratt R. Verkerk P. Zabel 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1997,94(1):115-122
The map positions of five previously described phytochrome genes have been determined in tomato (Lycopersicon esculentum Mill.) The position of the yg-2 gene on chromosome 12 has been confirmed and the classical map revised. The position of the phytochrome A (phy A)-deficient
fri mutants has been refined by revising the classical map of chromosome 10. The position of the PhyA gene is indistinguishable from that of the fri locus. The putative phyB1-deficient tri mutants were mapped by classical and RFLP analysis to chromosome 1. The PhyB1 gene, as predicted, was located at the same position. Several mutants with the high pigment (hp) phenotype, which exaggerates phytochrome responses, have been reported. Allelism tests confirmed that the hp-2 mutant is not allelic to other previously described hp (proposed here to be called hp-1) mutants and a second stronger hp-2 allele (hp-2
j
) was identified. The hp-2 gene was mapped to the classical, as well as the RFLP, map of chromosome 1.
Received: 24 May 1996 / Accepted: 14 June 1996 相似文献