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PubMed2003

Critical role of Ca2+ ions in the reaction mechanism of Euphorbia characias peroxidase.

A cationic peroxidase was isolated and characterized from the latex of the perennial Mediterranean plant Euphorbia characias. The purified enzyme contained one heme prosthetic group identified as ferric iron-protoporphyrin IX. In addition, the purified peroxidase contained 1 mol of endogenous calcium per mol of enzyme; removal of this calcium ion resulted in almost complete loss of the enzyme activity. However, when excess Ca(2+) was added to the native enzyme the catalytic efficiency was enhanced by 3 orders of magnitude. The mechanism of activation was studied using a wide range of spectroscopic and analytic techniques. Analysis of the steady state by stopped-flow measurements suggests that the main effect of calcium ions is to favor the oxidation of the ferric enzyme by hydrogen peroxide to form compound I, whereas the other steps of the catalytic cycle seem to be affected to a lesser extent. UV/vis absorption spectra and CD measurements show that the heme iron is pentacoordinated high-spin in native enzyme and remains so after the binding of Ca(2+). Only minor changes in the secondary or tertiary structure of the protein could be detected by fluorescence or CD measurements in the presence of Ca(2+) ions, except for a significant perturbation of the Fe(3+) inner sphere geometry, as detected by EPR measurements. We propose that Ca(2+) binding to a low affinity site induces a reorientation of the distal histidine changing the almost inactive form of Euphorbia peroxidase to a high activity form. This is the first example of a peroxidase that responds as an on/off switch to variations in the external Ca(2+) level.

Binding Sites↗
PubMed2005

Tryptophan-based radical in the catalytic mechanism of versatile peroxidase from Bjerkandera adusta.

Versatile peroxidase (VP) from Bjerkandera adusta is a structural hybrid between lignin (LiP) and manganese (MnP) peroxidase. This hybrid combines the catalytic properties of the two above peroxidases, being able to oxidize typical LiP and MnP substrates. The catalytic mechanism is that of classical peroxidases, where the substrate oxidation is carried out by a two-electron multistep reaction at the expense of hydrogen peroxide. Elucidation of the structures of intermediates in this process is crucial for understanding the mechanism of substrate oxidation. In this work, the reaction of H(2)O(2) with the enzyme in the absence of substrate has been investigated with electron paramagnetic resonance (EPR) spectroscopy. The results reveal an EPR signal with partially resolved hyperfine structure typical of an organic radical. The yield of this radical is approximately 30%. Progressive microwave power saturation measurements indicate that the radical is weakly coupled to a paramagnetic metal ion, suggesting an amino acid radical in moderate distance from the ferryl heme. A tryptophan radical was identified as a protein-based radical formed during the catalytic mechanism of VP from Bjerkandera adusta through X-band and high-field EPR measurements at 94 GHz, aided by computer simulations for both frequency bands. A close analysis of the theoretical model of the VP from Bjerkandera sp. shows the presence of a tryptophan residue near to the heme prosthetic group, which is solvent-exposed as in the case of LiP and other VPs. The catalytic role of this residue in a long-range electron-transfer pathway is discussed.

Basidiomycota↗
PubMed2004

Paracoccus pantotrophus pseudoazurin is an electron donor to cytochrome c peroxidase.

The gene for pseudoazurin was isolated from Paracoccus pantotrophus LMD 52.44 and expressed in a heterologous system with a yield of 54.3 mg of pure protein per liter of culture. The gene and protein were shown to be identical to those from P. pantotrophus LMD 82.5. The extinction coefficient of the protein was re-evaluated and was found to be 3.00 mM(-1) cm(-1) at 590 nm. It was confirmed that the oxidized protein is in a weak monomer/dimer equilibrium that is ionic-strength-dependent. The pseudoazurin was shown to be a highly active electron donor to cytochrome c peroxidase, and activity showed an ionic strength dependence consistent with an electrostatic interaction. The pseudoazurin has a very large dipole moment, the vector of which is positioned at the putative electron-transfer site, His81, and is conserved in this position across a wide range of blue copper proteins. Binding of the peroxidase to pseudoazurin causes perturbation of a set of NMR resonances associated with residues on the His81 face, including a ring of lysine residues. These lysines are associated with acidic residues just back from the rim, the resonances of which are also affected by binding to the peroxidase. We propose that these acidic residues moderate the electrostatic influence of the lysines and so ensure that specific charge interactions do not form across the interface with the peroxidase.

Azurin↗
PubMed2005

A Ca2+/calmodulin-binding peroxidase from Euphorbia latex: novel aspects of calcium-hydrogen peroxide cross-talk in the regulation of plant defenses.

Calmodulin (CaM) is a ubiquitous Ca(2+) sensor found in all eukaryotes, where it participates in the regulation of diverse calcium-dependent physiological processes. In response to fluctuations of the intracellular concentration of Ca(2+), CaM binds to a set of unrelated target proteins and modulates their activity. In plants, a growing number of CaM-binding proteins have been identified that apparently do not have a counterpart in animals. Some of these plant-specific Ca(2+)/CaM-activated proteins are known to tune the interaction between calcium and H(2)O(2) in orchestrating plant defenses against biotic and abiotic stresses. We previously characterized a calcium-dependent peroxidase isolated from the latex of the Mediterranean shrub Euphorbia characias (ELP) [Medda et al. (2003) Biochemistry 42, 8909-8918]. Here we report the cDNA nucleotide sequence of Euphorbia latex peroxidase, showing that the derived protein has two distinct amino acid sequences recognized as CaM-binding sites. The cDNA encoding for an E. characias CaM was also found and sequenced, and its protein product was detected in the latex. Results obtained from different CaM-binding assays and the determination of steady-state parameters showed unequivocally that ELP is a CaM-binding protein activated by the Ca(2+)/CaM system. To the best of our knowledge, this is the first example of a peroxidase regulated by this classic signal transduction mechanism. These findings suggest that peroxidase might be another node in the Ca(2+)/H(2)O(2)-mediated plant defense system, having both positive and negative effects in regulating H(2)O(2) homeostasis.

Amino Acid Sequence↗
PubMed2006

Catalase-peroxidase active site restructuring by a distant and "inactive" domain.

Catalase-peroxidases are composed of two peroxidase-like domains. The N-terminal domain contains the heme-dependent, bifunctional active site. The C-terminal domain does not bind heme, has no catalytic activity, and is separated from the active site by >30 A. Nevertheless, without the C-terminal domain, the N-terminal domain exhibits neither catalase nor peroxidase activity due to the apparent coordination of the distal histidine to the heme iron. Here we report the ability of the separately expressed and isolated C-terminal domain (KatG(C)) to restructure the N-terminal domain (KatG(N)) to its bifunctional conformation. Addition of equimolar KatG(C) to KatG(N) decreased the hexacoordinate low-spin heme complex and increased the high-spin species (pentacoordinate and hexacoordinate). EPR spectra of the domain mixture showed a distribution between high-spin species nearly identical to that of wild-type KatG. The CD spectrum for the 1:1 physical mixture of the domains was identical to an arithmetic composite of individual spectra for KatG(N) and KatG(C). Both physical and arithmetic mixtures were nearly identical to the spectrum for wild-type KatG, suggesting that major shifts in secondary structure did not accompany active site reconfiguration. With the shift in heme environment, the parallel return of catalase and peroxidase activity was observed. Inclusion of bovine serum albumin instead of KatG(C) produced no activity, indicating that specific interdomain interactions were required to reestablish the bifunctional active site. Apparent constants for reactivation (k(react) approximately 4 x 10(-3) min(-1)) indicate that a slow process like movement of established structural elements may precede the restructuring of the heme environment and return of catalytic activity.

Binding Sites↗
PubMed1996

Structure of the native cysteine-sulfenic acid redox center of enterococcal NADH peroxidase refined at 2.8 A resolution.

In order to obtain the crystal structure of the flavoprotein NADH peroxidase with its native Cys42-sulfenic acid redox center, a strategy combining reduced exposure of crystals to ambient oxygen and data collection at -160 degrees C was applied. The structure of the native enzyme to 2.8 A resolution is described; these results conclusively establish the existence of the Cys42-sulfenic acid as the functional non-flavin redox center of the peroxidase and provide the first structure for any naturally occurring protein-sulfenic acid. The Cys42-sulfenic acid atoms C alpha-C beta-S gamma-O roughly define a planar arrangement which is stacked parallel to the si face of the FAD isoalloxazine and positions the sulfenyl oxygen atom only 3.3 A from FAD-C4A. His10-N epsilon 2 contributes a hydrogen bond to the sulfenic acid oxygen, at a distance of 3.2 A. Although one oxygen atom (OX1) of the non-native Cys42-sulfonic acid derivative identified in the earlier wild-type peroxidase structure was taken to represent the native Cys42-sulfenic acid oxygen [Stehle, T., Ahmed, S. A., Claiborne, A., & Schulz, G. E. (1991) J. Mol. Biol. 221, 1325-1344], this structure shows that the sulfenic acid oxygen does not occupy this position, nor is it hydrogen-bonded to Cys42-N as was OX1. Comparison of the native Cys42-sulfenic acid structure with that of two-electron reduced glutathione reductase provides an insight into the sulfenic acid FAD charge-transfer interaction observed with both wild-type and His10 mutant peroxidases. A model of the E.NADH intermediate recently observed in stopped-flow analyses of the enzyme [Crane, E. J., III, Parsonage, D., Poole, L. B., & Claiborne, A. (1995) Biochemistry 34, 14114-14124] has also been generated to assist in analyzing the chemical mechanism of sulfenic acid reduction.

Amino Acid Sequence↗
PubMed1996

EPR detection and characterization of lignin peroxidase porphyrin pi-cation radical.

Lignin peroxidase (LiP) from Phanerochaete chrysosporium catalyzes the H2O2 dependent one- and two-electron oxidations of substrates. The catalytic cycle involves the oxidation of ferric-LiP by H2O2 by two electrons to compound I, which is an oxoferryl heme and a free radical. It has been speculated that the unpaired electron is in a pi delocalized porphyrin radical. However, no direct evidence for the presence of the free radical has been reported. We present electron paramagnetic resonance (EPR) detection and characterization of compound I of LiP. The LiP compound I EPR signal is different than those reported previously for compound I of horseradish peroxidase and chloroperoxidase. However, the EPR signal of compound I of LiP (axial g tensor extending from gperpendicular = 3.42 to gparallel approximately 2) is very similar to the EPR signals of compound I of ascorbate peroxidase and catalase from Micrococcus lysodeikticus, in which the radical has been identified as a porphyrin pi-cation radical. On the basis of the analysis of our data and comparison with the earlier published results for compounds I of other peroxidases, we interpret the LiP compound I signal by a model for exchange coupling between an S = 1 oxyferryl [Fe = O]2+ moiety and a porphyrin pi-cation radical (S = 1/2) [Schulz, C.E., et al. (1979) FEBS Lett. 103, 102-105]. The exchange coupling is characterized by ferromagnetic rather than an antiferromagnetic interaction between the two species. The ferric-Lip EPR signal suggests that the iron in the heme is in near perfect orthogonal symmetry and provides additional evidence of the ferromagnetic interaction between the oxoferryl iron center and the porphyrin pi-cation radical.

Basidiomycota↗
PubMed1997

pH dependence and structural interpretation of the reactions of Coprinus cinereus peroxidase with hydrogen peroxide, ferulic acid, and 2,2'-azinobis.

Steady-state and transient-state analysis of Coprinus cinereus peroxidase, CIP (identical to Arthromyces ramosus peroxidase), was used to characterize the kinetics of the three fundamental steps in heme peroxidase catalysis: compound I (cpd I) formation, cpd I reduction, and compound II (cpd II) reduction. The rate constant k1 for cpd I formation determined by transient-state analysis is (9.9 +/- 0.6) x 10(6) M-1 s-1. The k1 determined by steady-state analysis is (8.8 +/- 0.6) x 10(6) M-1 s-1 in the presence of ferulic acid and (6.7 +/- 0.2) x 10(6) M-1 s-1 in the presence of ABTS. The value of k1 is constant from pH 6 to 11. However, at low pH the value of k1 decreases, corresponding to titration of an enzyme group with a pKa of 5.0. Titration of this group is also detected from cyanide-binding kinetics. Furthermore, titration of this group is linked with marked spectroscopic changes unique to CIP. We ascribe these changes to protonation of proximal His183. A very low pKa is proposed for distal His55 in the resting state of CIP. The rate constants, k2 for cpd I and k3 for cpd II reduction, are very large for both ferulic acid and 2,2'-azinobis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS). For ferulic acid, transient-state kinetic analysis shows that the values of k2 and k3 are identical at pH 5-6, and the ratio k2/k3 increases to 10 at pH 10. The similar magnitude of k2 and k3 is unusual for a peroxidase. Both k2 and k3 decrease with increasing pH, and both are influenced by two ionizations: one with a pKa value near 7, assumed to reflect the protonation of His55; and the other with pKa of 9.0 +/- 0.7 for k2 and 8.8 +/- 0.4 for k3, perhaps reflecting the phenol-linked deprotonation of ferulic acid. Steady-state analysis at pH 7.0 gave k2k3/(k2 + k3) = (2.2 +/- 0.1) x 10(7) M-1 s-1 for ferulic acid, and (2.0 +/- 0.7) x 10(7) M-1 s-1 for ABTS and revealed a unimolecular step with ku = 1500 s-1, ascribed to slow ABTS radical product release. From transient-state results at pH 7, the values of k2 and k3 were found to be identical also for ABTS. A mechanism for cpd I and II reduction involving distal histidine and arginine is proposed.

Benzothiazoles↗
PubMed1997

Detection and characterization of the lignin peroxidase compound II-veratryl alcohol cation radical complex.

Lignin peroxidases (LiP) from the white-rot fungus Phanerochaete chrysosporium oxidize veratryl alcohol (VA) by two electrons to veratryl aldehyde, although the VA cation radical (VA.+) is an intermediate [Khindaria, A., et al. (1995) Biochemistry 34, 6020-6025]. It was speculated, on the basis of kinetic evidence, that VA*+ can form a catalytic complex with LiP compound II. We have used low-temperature EPR to provide direct evidence for the formation of the complex. The EPR spectrum of VA*+ obtained at 4 K was explained by a model for coupling between the oxoferryl moiety of the heme (S = 1) and VA.+ (S = 1/2) similar to the model proposed for an oxyferryl and a porphyrin pi cation radical of horseradish peroxidase. The coupling constant suggested that VA.+ was equally ferro- and antiferromagnetically coupled to the oxoferryl moiety. The spectrum was simulated with g perpendicular only marginally greater than g parallel. This was surprising since the only other known organic radical coupled to the heme iron in a peroxidase is the tryptophan cation radical in cytochrome c peroxidase which exhibits a g tensor with g parallel greater than g perpendicular. Spin concentration analysis suggested that the 1 mol of VA*+ was coupled to the oxoferryl moiety per mole of enzyme. The VA.+ signal decayed with a first-order decay constant of 1.76 s-1, in close agreement with the earlier published decay constant of 1.85 s-1 from room-temperature EPR studies. The exchange coupling between VA.+ and the oxoferryl moiety strongly advocates calling this species (VA.+ and LiP compound II) a catalytic complex.

Basidiomycota↗
PubMed1998

The role of naturally occurring phenols in inducing oscillations in the peroxidase-oxidase reaction.

The influence of a series of naturally occurring phenolic compounds on the dynamics of the horseradish peroxidase-catalyzed oxidation of NADH by oxygen (the peroxidase-oxidase reaction) was investigated. Various types of dynamic behaviors are induced in the peroxidase-oxidase reaction upon addition of low concentrations of different phenols. The identity of the particular phenol determines the type of dynamics shown (inhibition of the reaction, monotonic reaction, damped oscillations, sustained oscillations, and complex oscillations). The kind of behavior that a given phenolic compound is able to induce is governed by the reduction potentials for the formation of the phenoxyl radical. The phenolic compounds are shown not to be consumed during the reaction. Mechanistic considerations are made on the basis of the reduction potentials of the different reactants present in the peroxidase-oxidase system. We interpret the role of the phenols as electron mediators of the electron transfer between some enzyme intermediates or O2- and NADH.

Electrochemistry↗
PubMed1998

Structural roles of the highly conserved glu residue in the heme distal site of peroxidases.

One of the highly conserved amino acid residues in the heme distal site of various fungal and plant peroxidases, glutamic acid 64 (Glu64) in horseradish peroxidase (HRP), interacts with a distal calcium ion through a hydrogen bond with a water molecule and its peptide carbonyl oxygen on the main-chain forms the hydrogen bond network to the distal His via the adjacent Asn residue, suggesting that the Glu residue is related to the stabilization of the calcium ion and catalytic activity of peroxidase [Nagano, S., Tanaka, M., Ishimori, K., Watanabe, Y., and Morishima, I. (1996) Biochemistry 35, 14251-14258]. To perturb the hydrogen bond with the adjacent Asn, we replaced the Glu with Pro (E64P) or Gly (E64G), which would alter the configuration of the main chain at position 64. Both of the mutants exhibited substantially depressed oxidation activities for hydroquinone and elementary reaction rates in the catalytic cycle. However, the E64S (Glu64 --> Ser) mutant, in which the configuration of the main chain and the hydrogen bond with Asn70 would not be affected but the interactions with the calcium ion are seriously perturbed by removal of the carboxylate, also showed quite low catalytic activity as observed for the E64P and E64G mutants. Spectral features for the E64S mutant are similar to those of the other mutants: the reorientation of the distal His, disruption of the hydrogen bond between the distal His and Asn70, and loss of the calcium ion. Thus, we can conclude that, in addition to forming the hydrogen bond network in the distal site, the Glu residue is a key residue for stable binding of the calcium ion, which maintains the structural integrity of the distal cavity, resulting in high peroxidase activity.

Amino Acid Sequence↗
PubMed1998

The distal cavity structure of carbonyl horseradish peroxidase as probed by the resonance Raman spectra of His 42 Leu and Arg 38 Leu mutants.

CO ligation to horseradish peroxidase C (HRPC) was studied by means of site-directed mutagenesis and resonance Raman spectroscopy. The CO complexes of HRPC His 42 --> Leu and Arg 38 --> Leu mutants were characterized at pH values ranging from 3.6 to 9.5. The vibrational frequencies of the Fe-C stretching and Fe-C-O bending modes have been identified by isotopic substitution. Both His 42 --> Leu and Arg 38 --> Leu adducts with CO displayed a single Fe-C stretching band, whereas both recombinant and wild-type HRPC-CO have two bands, corresponding to different conformers. This comparison suggests that CO is H-bonded either to the distal Arg or to the distal His in the two conformers. An acid transition, common to the wild-type protein, was observed for both mutants. This indicates that these distal amino acids do not influence the acid transition. On the contrary, an alkaline transition was only observed for the Arg 38 --> Leu mutant, which suggests that distal His is involved in the alkaline transition of HRPC-CO complex. The spectroscopic information is found to be consistent with the X-ray structure of ferric HRPC. A comparison with the CO complexes of cytochrome c peroxidase and myoglobin is performed, which displays the functional significance of the structural differences between peroxidase classes I and III and between peroxidases and globins, respectively.

Amino Acid Substitution↗
PubMed1999

Binding of salicylhydroxamic acid and several aromatic donor molecules to Arthromyces ramosus peroxidase, investigated by X-ray crystallography, optical difference spectroscopy, NMR relaxation, molecular dynamics, and kinetics.

The X-ray crystal structure of the complex of salicylhydroxamic acid (SHA) with Arthromyces ramosus peroxidase (ARP) has been determined at 1.9 A resolution. The position of SHA in the active site of ARP is similar to that of the complex of benzhydroxamic acid (BHA) with ARP [Itakura, H., et al. (1997) FEBS Lett. 412, 107-110]. The aromatic ring of SHA binds to a hydrophobic region at the opening of the distal pocket, and the hydroxamic acid moiety forms hydrogen bonds with the His56, Arg52, and Pro154 residues but is not asscoiated with the heme iron. X-ray analyses of ARP-resorcinol and ARP-p-cresol complexes failed to identify the aromatic donor molecules, most likely due to the very low affinities of these aromatic donors for ARP. Therefore, we examined the locations of these and other aromatic donors on ARP by the molecular dynamics method and found that the benzene rings are trapped similarly by hydrophobic interactions with the Ala92, Pro156, Leu192, and Phe230 residues at the entrance of the heme pocket, but the dihedral angles between the benzene rings and the heme plane vary from donor to donor. The distances between the heme iron and protons of SHA and resorcinol are similar to those obtained by NMR relaxation. Although SHA and BHA are usually considered potent inhibitors for peroxidase, they were found to reduce compound I and compound II of ARP and horseradish peroxidase C in the same manner as p-cresol and resorcinol. The aforementioned spatial relationships of these aromatic donors to the heme iron in ARP are discussed with respect to the quantum chemical mechanism of electron transfer in peroxidase reactions.

Binding Sites↗
PubMedDate not supplied

Synthesis of conducting polyelectrolyte complexes of polyaniline and poly(2-acrylamido-3-methyl-1-propanesulfonic acid) catalyzed by pH-stable palm tree peroxidase.

Comparison of the stability of five plant peroxidases (horseradish, royal palm tree leaf, soybean, and cationic and anionic peanut peroxidases) was carried out under acidic conditions favorable for synthesis of polyelectrolyte complexes of polyaniline (PANI). It demonstrates that palm tree peroxidase has the highest stability. Using this peroxidase as a catalyst, the enzymatic synthesis of polyelectrolyte complexes of PANI and poly(2-acrylamido-3-methyl-1-propanesulfonic acid) (PAMPS) was developed. The template polymerization of aniline was carried out in aqueous buffer at pH 2.8. Varying the concentrations of aniline, PAMPS, and hydrogen peroxide as reagents, favorable conditions for production of PANI were determined. UV-vis-NIR absorption and EPR demonstrated that PAMPS and PANI formed the electroactive complex similar to PANI doped traditionally using low molecular weight sulfonic acids. The effect of pH on conformational variability of the complex was evaluated by UV-vis spectroscopy. Atomic force microscopy showed that a size of the particles of the PANI-PAMPS complexes varied between 10 and 25 nm, depending on a concentration of PAMPS in the complex. The dc conductivity of the complexes depends also on the content of PAMPS, the higher conductivity being for the complexes containing the lower content of the polymeric template.

Aniline Compounds↗
PubMedDate not supplied

2,4-Dichlorophenol degradation using Streptomyces viridosporus T7A lignin peroxidase.

The Streptomyces viridosporus T7A bacterium produces the extracellular lignin peroxidase ALiP-P3. The ALiP-P3-catalyzed oxidation of 2,4-dichlorophenol (DCP) was examined to understand its kinetic behavior. Initial rate data of the oxidation of DCP were obtained by a spectrophotometric peroxidase assay, and the kinetics were best modeled with a random-binding bireactant system, which differs from the ping-pong bireactant system that is typically used for horseradish peroxidase and lignin peroxidase from the fungus Phanerochaete chrysosporium, and suggests that either DCP or H2O2 may bind first to ALiP-P3. Chloride ion measurements indicate that 16% of the reacted DCP was fully dechlorinated by ALiP-P3. Chemical ionization mass spectrometry was also utilized to identify the DCP degradation product as a hydrophobic chlorinated dimer of mass 322.

Anions↗
PubMed2001

Glutathione peroxidase-like antioxidant activity of diaryl diselenides: a mechanistic study.

The synthesis, structure, and thiol peroxidase-like antioxidant activities of several diaryl diselenides having intramolecularly coordinating amino groups are described. The diselenides derived from enantiomerically pure R-(+)- and S-(-)-N,N-dimethyl(1-ferrocenylethyl)amine show excellent peroxidase activity. To investigate the mechanistic role of various organoselenium intermediates, a detailed in situ characterization of the intermediates has been carried out by (77)Se NMR spectroscopy. While most of the diselenides exert their peroxidase activity via selenol, selenenic acid, and selenenyl sulfide intermediates, the differences in the relative activities of the diselenides are due to the varying degree of intramolecular Se.N interaction. The diselenides having strong Se.N interactions are found to be inactive due to the ability of their selenenyl sulfide derivatives to enhance the reverse GPx cycle (RSeSR + H(2)O(2) = RSeOH). In these cases, the nucleophilic attack of thiol takes place preferentially at selenium rather than sulfur and this reduces the formation of selenol by terminating the forward reaction. On the other hand, the diselenides having weak Se.N interactions are found to be more active due to the fast reaction of the selenenyl sulfide derivatives with thiol to produce diphenyl disulfide and the expected selenol (RSeSR + PhSH = PhSSPh + RSeH). The unsubstituted diaryl diselenides are found to be less active due to the slow reactions of these diselenides with thiol and hydrogen peroxide and also due to the instability of the intermediates. The catalytic cycles of 18 and 19 strongly resemble the mechanism by which the natural enzyme, glutathione peroxidase, catalyzes the reduction of hydroperoxides.

Antioxidants↗
PubMed2003

Retention and extractability of phenol, cresol, and dichlorophenol exposed to two surface soils in the presence of horseradish peroxidase enzyme.

The retention of phenol, o-cresol, 2,4-dichlorophenol (DCP), and their peroxidase-catalyzed polymerization products was evaluated on two surface soils. The extractability of the parent solutes and their polymerization products was also investigated. (14)C-Labeled radioisotopes were used to quantify the contaminant retained on soil as water-extractable, methanol-extractable, humic/fulvic (HA/FA) acid-bound, and soil/humin bound. Between 2 and 20% of the solute retained on soil after a 7-day contact period remained bound to the HA/FA and soil/humin components in unamended soils; in the presence of peroxidase this amount was as high as 40-75%. The alkali-extractable HA/FA component contained the largest fraction of radioactivity in peroxidase-amended soils. Whereas the soil organic matter content was the predominant factor controlling the extent of sorption of the parent phenols, the clay content and particle surface area appeared to contribute to the retention of the polymerization products. High molecular weight oligomers produced during peroxidase-mediated polymerization of phenols associate strongly with soil components and are likely incorporated into the soil organic matter via oxidative coupling reactions.

Benzopyrans↗
PubMed2000

Purification and partial characterization of three turnip (Brassicanapus L. var. esculenta D.C.) peroxidases.

Three turnip peroxidases (fractions C1, C2, and C3) were partially purified and characterized, to permit study of their feasibility for use in clinical and enzyme immunoassays. These fractions represented 20% of the initial activity, and fractions C1 and C2 were purified to homogeneity. The optimum pH was between 5.0 and 5.5, while optimum temperature ranged from 40 to 55 degrees C. The ABTS K(m) values for the two acidic fractions (C2 and C3) were 0.70 and 0.42 mM, respectively; about 5 times lower than that reported for the acidic commercial horseradish peroxidase (HRP). Fraction C3 had 4 times higher K(m) value than commercial cationic HRP. The molecular weights determined by SDS-PAGE ranged from 39.2 to 42.5 kDa. Activation energies for inactivation were 113 (C1), 130 (C2), and 172 kJ/mol (C3) which are higher or comparable to other peroxidase isoenzymes reported. Fractions C1 and C3 represent an alternative source of peroxidase because of their higher purification yield and specific activity, when compared to fraction C2.

Brassica↗