Peroxidases isolation

Attachment of Hydroxycinnamic Acids to Structural Cell Wall Polymers. Peroxidase mediation may also result in binding the hydroxycinnamic acids to the plant cell wall polymers (66,67). For example, it was reported that peroxidases isolated from the cell walls of Pinus elliottii catalyze the formation of alkali-stable linkages between [2-14C] ferulic acid 1 and pine cell walls (66). Presumably this is a consequence of free-radical coupling of the phenoxy radical species (from ferulic acid 1) with other free-radical moieties on the lignin polymer. There is some additional indirect support for this hypothesis, since we have established that E-ferulic acid 1 is a good substrate for horseradish peroxidase with an apparent Km (77 /tM), which is approximately one fifth of that for E-coniferyl alcohol (400 /iM) (unpublished data). 

However, it should be noted that isoenzymes, i.e., different enzymes catalyzing identical reactions, would have the same four-digit classification. The classification system provides only the basis for a unique identification of an enzyme the particular isoenzyme and its source still have to be specified. For example, peroxidases isolated from soybeans and horseradishes have the same classification, i.e., EC 1.11.1.7. Currently, there are approximately 3200 enzymes that have been listed and assigned classification numbers. 

Ruiz-Duenas FJ, Martinez MJ, Martinez AT (1999) Molecular characterization of a novel peroxidase isolated from the ligninolytic fungus Pleurotus eryngii. Mol Microbiol 31 223-235... 

S. Braddon-Galloway, J. E. Balthrop, Se-dependent GSH-peroxidase isolated from Black Sea Bass (Centropristis striata), Comp. Biochem. Physiol., 82C (1985), 297-300. 

The manganese(III)-phenoxide couple is quite unstable in organic chemistry, Mn(III) and Mn(IV) complexes have been demonstrated to be excellent oxidants for phenols and their analogs (145). Indeed, a peroxidase isolated from white rot fungus is dependent on extracellular Mn(II) (146-149). The heme-peroxide moiety of this enzyme is reduced by the Mn, which in turn (as the trivalent species) migrates to the phenolic substrate. The substrate is thus oxidized by the generated... 

Class III peroxidases isolated from C. roseus are capable of oxidizing ajmalicine (XLVIII) to serpentine (LV) [150]. The kz value of C. roseus peroxidase for this reaction is about 0.0011 pM s at pH 6.0 [74]. This constitutes a unique example in peroxidase-catalyzed metabolic reactions since it involves the aromatization of a N-heterocyclic ring (Scheme XXV). 

There is a third class of peroxidases isolable from the bacterial sources such as P. aeruginosa and Pseudomonas stutzerii, which oxidize cytochrome C551, or azurin. This protein contains two heme protoporphyrin IX groups covalently bound to a single polypeptide side chain. In this enzyme one heme group is oxidized from Fe(III) to Fe(IV)=0 and the second heme, from Fe(II) to Fe(III). The oxidizing equivalents are directed to two centers with very different redox potentials (75). 

Two different catalysts for hydrogen peroxide decomposition, the enzyme peroxidase (isolated from the horseradish root, HRP), and polymer-supported catalyst (acid form of poly-4-vinylpyri-dine functionalized by ferric sulfate, apFe) [99,100], are examined with an aim to compare their activity. The active center in the peroxidases is the ferric ion in protoporphyrin IX. Besides the complex made of ferric ion and protoporphyrin IX, that is ferricprotoporphyrin IX, also known as ferric heme or hemin, peroxidase possesses a long chain of proteins [101,102]. On the other hand, the macroporous acid form of polyvinyl pyridine functionalized by ferricsulfate is obtained from cross-linked polyvinyl pyridine in macroporous bead form [103]. Pyridine enables it to form coordination complexes or quaternary salts with different metal ions such as iron (111) [104]. An active center on the polymeric matrix functionalized by iron, as metallic catalyst immobilized on polymer by pyridine, has similar microenvironment conditions as active center in an enzyme [105]. 

Oh, S. H., Ganther, H. E., and Hoekstra, W. G., 1974, Selenium as a component of glutathione peroxidase isolated from ovine erythrocytes. Biochemistry 13 1825. 

Glutathione peroxidase isolated from erythrocytes has a relative molecular weight of 85-95 kDa and consists of four subunits, each of which contains one selenocysteine molecule in its peptide chain. 

Thermal Inactivation of Peroxidase. To study thermostability of spinach peroxidase without the influence of cellular components, isolated peroxidase was heat treated in 0.1 M phosphate buffer (pH 6.0). The thermal inactivation curves are presented in Figure 6 and they showed biphasic kinetic curves m the range of 60-70 C. The spinach suspension and the extract (Figure 6b and c) also exhibited biphasic curves similar to that of isolated peroxidase (Figure 6a). The thermodynamic data were siunmarized in Table m. The results indicated that isolated peroxidase was more thermostable than those in the suspension or in the extract in the range of 50-80 t). However, z-value was 13 t) for isolated peroxidase which was less than those ( 18 TO) for both of the enzymes in the suspension or tiie extract The results showed that peroxidase isolated fix>m spinach responded differently to heating than the enzymes in the spinach extract or suspension. In Table IV, heat stabilities of peroxi-... 

Other specific discovery assays have been used such as differential inhibition of a vancomycin resistant S. aureus strain and its susceptible parent, and an assay based on antagonism of the antibacterial activity by N,A/-diacetyl-L-Lys-D-Ala-D-Ala [24570-39-6] a tripeptide analogue of the dalbaheptides receptor. AppHcation of this latter test to 1936 cultures (90) led to the isolation of 42 dalbaheptides, six of which, including kibdelin (Table 3), parvodicin (Table 3), and actinoidin A2 (68) were novel. A colorimetric assay based on competition between horseradish peroxidase bound teicoplanin and the... 

Enzyme-mediated chiral sulfoxidation has been reviewed comprehensively in historical context [188-191]. The biotransformation can be mediated by cytochrome P-450 and flavin-dependent MOs, peroxidases, and haloperoxidases. Owing to limited stability and troublesome protein isolation, a majority of biotransformations were reported using whole-cells or crude preparations. In particular, fungi have been identified as valuable sources of such biocatalysts and the catalytic entities have not been fully identified in all cases. 

Yang Q, Yang M, Pritsch K et al (2003) Decolorization of synthetic dyes and production of manganese-dependent peroxidase by new fungal isolates. Biotechnol Lett 25 709-713... 

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