Peroxidase compounds, conversion

Compounds I and II are quite stable at low temf>erature and therefore can serve as pure reactants to study the mechanisms of peroxide oxidase processes (Douzou, 197la,b). When compound II reacted with indole 3-acetate, this compound was immediately regenerated without the appearance of any other intermediary compound. Moreover, indole 3-acetate in large excess induced the conversion of compound III into compound II. A study of reaction mechanisms of indole 3-acetate degradation by various peroxidases was recently carried out by Ricard and Job (1974) using low-temperature spectroscopic techniques. They obtained new data that made it possible to propose electronic mechanisms of reactions less speculative than those dependent upon data obtained under normal conditions of temperature. 

Barr DP, Aust SD (1994) Conversion of lignin peroxidase compound III to active enzyme by cation radicals. Arch Biochem Biophys 312 511-515... 

Comparison of the rate constants of [(TMP)Fe =0] + and HRP compound I was further extended to reactions with a series of sulfides [108]. HRP is known to convert thioanisole to the corresponding sulfoxide [109], although peroxidases typically catalyze two sequential one-electron oxidations such as one-electron oxidation of phenol derivatives to phenoxy radicals [110], The yield of the sulfoxide from the stoichiometric reaction of HRP compound I with thioanisole is only 25 + 5 %. The sulfoxidation involves oxygen-transfer from an oxoferryl species to sulfide, because in H2 02 has been shown to be incorporated into the product sulfoxide [108, 111], The initial rapid conversion of compound I to compound II (fci) by thioanisole is followed by further reduction to the ferric resting state, as is found for reactions with DMA [108, 112, 113]. A linear correlation between log A i and E°ox for the reactions of HRP compound I with thioanisoles (Figure 2) is readily combined with the relationship for DMA (Figure la) into a single, common relationship (Eq. 8) [108]. 

Figure 18. Reaction scheme for conversion of peroxidase compounds...

LiP catalyzes the oxidation of 3,4-dimethoxybenzyl alcohol (veratryl alcohol) to veratryl aldehyde. Since this reaction can be easily followed at 310 nm, it is the basis for the standard assay for this enzyme (26,27). The enzyme exhibits normal saturation kinetics for both veratryl alcohol and H202 (28,43). Steady-state kinetic results Indicate a ping-pong mechanism in which H202 first oxidizes the enzyme and the oxidized intermediate reacts with veratryl alcohol (43). The enzyme has an extremely low pH optimum ( 2.5) for a peroxidase (43,44) however, the rate of formation of compound I (kx, Fig. 2) exhibits no pH dependence from 3.0-7.0 (45,46). Addition of excess veratryl alcohol at pH 3.0 results in the rapid conversion of... 

One might hope to see evidence in the 3D structure for both the ferric-ferryl conversion and the amino-acid free radical. However, X-ray crystallography of proteins cannot resolve individual hydrogen atoms. Therefore it is important to remember [104] that as the oxidation of cytochrome c peroxidase to compound I involves no change in the number of non-hydrogen atoms, it is possible that... 

Once Compound I has catalyzed the one-electron oxidation of a substrate, the porphyrin cation radical is "neutralized" and the so-called Compound II is formed (Fig. 4.77). This Compound n, perhaps upon protonation of its 0x0 atom, is still a powerful oxidizing species, and can catalyze the one electron oxidation of a second substrate molecule, leading also to formation of a molecule of water and conversion of the heme moiety to its Fe " resting state. Because the oxidative power of Compoimd II is lower than that of Compound I, substrate oxidation by Compound II is the rate-limiting step in peroxidase biocatalysis. 

The conversion of the green primary complex into the pale red secondary complex appears to be a reduction process even though it occurs in the absence of any added hydrogen donors. The most definite evidence for this is the case of peroxidase where the speed of the conversion is increased in the presence of all compounds with which the peroxide system reacts (Chance, 55). For catalase, where the conversion can only be obtained with alkyl hydroperoxides, the evidence is not so clear-cut, but at least the velocity of formation of the secondary complexes increases as the hydroperoxide concentration is increased. An alternative explanation for these effects would be that the primary and secondary complexes are in some sort of equilibrium where removal of the latter would have the effect of increasing the rate of conversion. There is no indication of any such equilibrium, however, and direct reduction of the primary complex appears to be the most likely explanation. One possible formulation for this change involves the production of a ferryl ion type of compound by the removal of an OH radical by the hydrogen donor from the 02H anion bound to the iron atom ... 

Hexanal phenylhydrazone also serves as an inactivator of soybean lipoxygenase 1 (L-1) in a process which demonstrates kinetics more complex than those of standard mechanism-based inactivators (Galey et al., 1988). Aerobic incubation of hexanal phenylhydrazone with L-1 leads to enzyme inactivation and conversion of the compound to its corresponding a-azo hydroperoxide, which is also an inactivator. Four equivalents of the a-azo hydroperoxide are sufficient to inactivate the enzyme completely, whereas the amount of the parent phenylhydrazone required to fiilly inactivate the enzyme increases from 13 to a maximum of 70 as the ratio of hexanal phenylhydrazone to L-1 increases. Since the partition ratio is normally independent of inhibitor and enzyme concentrations, a more complex mechanism is apparent. The addition to reaction mixtures of glutathione peroxidase, which reduces the a-azo hydroperoxide metabolite to the corresponding alcohol, suppresses about 80% of the inactivation. The a-azo hydro-... 

An alternative or complimentary theory for the mode of action of simple phenolic compounds is that they are converted to much more toxic quinones. Pillinger et al. [69] found that various phenolic decomposition products of barley straw were most toxic under conditions favorable for oxidation of the compounds to quinones, and that quinones were up to one thousand-fold more toxic to algae than the parent compounds. The most likely route to conversion to a quinone is enzymatic. Peroxidases and polyphenol oxidases can perform such a reaction, depending on the substrate. However, polyphenol oxidase cannot be detected in most green algae [139] and has not been reported in cyanobacteria. 

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