Role of Peroxidase in Lignification of Tobacco Cells

Abstract

Coniferyl alcohol is the prinary substrate for peroxidase-mediated ligniflcatlon, a process which depends on the generation of H202 by NADH oxidation. We measured the concentrations of various phenols (synthetic and natural) The precursors of lignin formation are generally thought to be the cinnamyl alcohols. The polymerization of these phenols is catalyzed by peroxidase in the presence of H202 (2, 3). The formation of H202 is also catalyzed by peroxidase resulting from an oxidation ofNADH (2, 7). We have studied the role ofdifferent isoenzyme groups of tobacco peroxidase in both reactions (6,9). In this paper, we report on the role of coniferylalcohol in the H202-forming reaction (NADH-oxidation) and compare it to other natural and synthetic phenols. The role of phenols is discussed with respect to their regulation of H202-formation and lignification in-the cell wall. MATERIALS AND METHODSAll materials and methods have been described in the accompanying (5) and in a previous paper (6). RESULTS pH-Dependence of NADH-Oxidation. Determination of the pH optimum for oxidation of NADH with DCP2 as cofactor with tobacco peroxidase isoenzyme group GI (electrophoretically fast migrating, anodic) and Gm (electrophoretically slow migrating, cathodic) indicated a value of 4.5. There is a second pH optimum ' Supported by the Deutsche Forschungsgemeinschaft.2Abbreviations: DCP, dichlorophenol; HRP, horseradish peroxidase. of 7.0 and 6.0 for GI and Gm, respectively, although the reaction rates are 30%o to 50%o lower (data not shown). With coniferyl alcohol as cofactor, only one pH optimum at 4.5 was found. These findings are in contrast to those obtained earlier in which H202 formation was determined by the ['4CJglyoxylate method (2, 9). However, with this method in which the nonenzymic oxidation of glyoxylate is coupled to the enzymic oxidation of NADH, the measured pH optimum is not that of NADH oxidation alone (9). With HRP, a pH optimum of 4.5 has also been found when NADH oxidation itself was measured.Phenols as Cofactors of NADH-Oxidation. It has been shown that maximal NADH oxidation occurs in the presence of both Mn2' and phenols (4)'.4Mn2' alone enhanced the rate of the reaction catalyzed by tobacco peroxidase 2-to 3-fold; phenols alone enhance the reaction rate 10-to 100-fold, but together they accelerate the oxidation of NADH 103-to 104-fold (5).Experiments were carried out to find the extent to which synthetic and naturally occurring phenols enhance the rate of NADH oxidation, and at which concentration the phenols must be present. The data (Table I and II) are expressed as a ratio of the rate of NADH oxidation (AAs4o) over the rate of tetraguaiacol oxidation (AA470). The oxidation of tetraguaiacol is thereby used as an internal standard allowing us to compare the effects of different phenols. The concentration necessary for the maximal rate of NADH oxidation was generally much higher (102_ to 104-fold) for synthetic phenols than for natural phenols. The data show that synthetic phenols which have the ...

The Role of Hydrogen Peroxide-Producing and Hydrogen Peroxide-Consuming Peroxidases in the Leaf Apoplast of Cowpea in Manganese Tolerance

“…The phenolic compounds had inconsistent effects on the functionality of PODs (Lee, 1977;Mäder and Fü ssl, 1982;Pedreñ o et al, 1987). Already Kenten and Mann (1950) detected a significant influence of several phenolic compounds on the POD-catalyzed oxidation of manganous pyrophosphate.…”

“…Mn oxidation was stimulated by monohydroxy phenols but not by dihydroxy and trihydroxy phenols. Phenols have been proposed to affect the peroxidase-catalyzed NADH oxidation via different pathways (Yamazaki and Yokota, 1973;Halliwell, 1978;Mäder and Fü ssl, 1982;Pedreñ o et al, 1987). The time course of NADH oxidation showed a characteristic lag time, followed by a high turnover rate of NADH (Yokota and Yamazaki, 1977;Pedreñ o et al, 1987).…”

“…The regulatory effect of phenolic compounds and Mn on NADH-peroxidase activity is widely documented (Yamazaki and Piette, 1963;Halliwell, 1978;Mäder et al, 1980;Mäder and Fü ssl, 1982;Pedreñ o et al, 1987). Mn II was proposed to affect the NADH-peroxidase cycle by reducing reactive oxygen, thus forming Mn III (Yamazaki and Piette, 1963;Halliwell, 1978).…”

“…Peroxidases using NADH as electron donor were also termed NADH oxidases and have been suggested to play an important role in the formation of H 2 O 2 needed for lignification (Elstner and Heupel, 1976;Gross et al, 1977;Halliwell, 1978;Mäder et al, 1980;Mäder and Amberg-Fisher, 1982;Bolwell et al, 1995). Mn and phenolic compounds presumably control the activity of the NADH peroxidase (Elstner and Heupel, 1976;Gross et al, 1977;Halliwell, 1978;Mäder and Fü ssl, 1982;Stich and Ebermann, 1984;Pedreñ o et al, 1987). Proteins purified from the apoplastic washing fluid (AWF) of leaves from the Mnsensitive cowpea cv TVu 91 showed NADH-peroxidase activity, which was strongly enhanced at high leaf Mn concentrations (Fecht-Christoffers et al, 2003a).…”

Biochemical characterization of Santalum album (Chandan) leaf peroxidase

“…In the presence of NADH (Yamazaki and Yokota 1973). In the peroxidatic reaction, phenolic compounds are especially effective as electron donors for the intermediates compound I (Fe 5+ ) and compound II (Fe 4+ ) (Mader and Fussl 1982).…”