Oxidase Reactions of Tomato Anionic Peroxidase

Abstract

ABSTRACITomato (Lycopersicon esculentum Mill) anionic peroxidase was found to catalyze oxidase reactions with NADH, glutathione, dithiothreitol, oxaloacetate, and hydroquinone as substrates with a mean activity 30% that of horseradish peroxidase; this is in contrast to the negligible activity of the tomato enzyme as compared to the horseradish enzyme in catalyzing an indoleacetic acid-oxidase reaction with only Mn2' and a phenol as cofactors. Substitution of Ce3' for Mn2' produced an 18-fold larger response with the tomato enzyme than with the horseradish enzyme, suggesting a significant difference in the autocatalytic indoleacetic acidoxidase reactions with these two enzymes. In attempting to compare enzyme activities with 2,4-dichlorophenol as a cofactor, it was found that reaction rates increased exponentially with both increasing cofactor concentration and increasing enzyme concentration. While the former response may be analogous to allosteric control of enzyme activity, the latter response is contrary to the principle that reaction rate is proportional to enzyme concentration, and additionally makes any comparison of enzyme activity difficult.Tomato ripening, like that of pears, bananas, and avocadoes, appears to be at least partly controlled by auxin levels (12); free IAA decreases and peroxides accumulate in tomato ripening.An isoelectric point of less than 5.5 for tomato anionic peroxidase, as suggested by its column chromatographic behavior, is lower than those of the major horseradish peroxidase isozymes (1 1) and is in particular lower than the value of 8.9 for horseradish peroxidase isozyme C, which predominates in commercial horseradish peroxidase preparations. This is of interest because of the statement (2) that cationic peroxidases serve as IAAoxidases in vivo. The more cationic members of isozyme sets often show the highest specific activity in the IAA-oxidase reaction or the highest ratio of IAA-oxidase to peroxidase activity (1 1, 20, 22, 24), though this is not consistently the case (5, 1 1). In considering the results of in vitro comparisons of peroxidases, it should be recalled that IAA-oxidase activity is more closely related to the oxidation-reduction potential of the enzyme than to the isoelectric point (24), that peroxidase isozymes may differ greatly in their peroxidase activity (15), and that relative IAAoxidase activities may depend greatly on the conditions of assay used (1, 22).The major peroxidase oftomato fruits, termed tomato anionic peroxidase, differs markedly from horseradish peroxidase in its ability to oxidize IAA (17). The horseradish enzyme catalyzes an autocatalytic IAA-oxidase2 reaction, which is stimulated by phenols and Mn2". The IAA-oxidase reaction of the tomato enzyme, however, was found to be completely dependent on the presence of catalytic amounts of H202 in the assay mixtures (18) MATERIALS AND METHODS Enzymes. Tomato anionic peroxidase was prepared from tomato pericarp using extraction, (NH4)2SO4 fractionation, and QAE-Sephadex column chromatography with grad...

Oxidation of Indoleacetic Acid by an Apparently Homogeneous Peroxidase From the Flavedo of Washington Navel Oranges (Citrus Sinensis)

“…The apparently conflicting results obtained by Kokkinakis and Brooks (1979a,b) and Brooks (1986) and by Thomas and Jen (1980), in relationship to the effect of H,O, on IAA oxidation by a tomato fruit peroxidase, could probably be explained in light of our results since, the former, utilized citrate and succinate buffer, at pH 4, while the latter used phosphate buffer at pH 6. Such different conditions could favor one of the reactions but be inadequate for the other one.…”

“…Such different conditions could favor one of the reactions but be inadequate for the other one. The experiments by Huang and Haard (1977) and Brooks (1986), with tomato fruit peroxidase extracts, seem to indicate a distinct effect of pH on IAA oxidase activity depending on the addition of H,02 to the reaction medium.…”

Consequences of chilling stress on watermelon [Citrullus lanatus (Thunb.) Matsum. and Nakai] germplasm lines at seedling stage

“…The primary function of the peroxidase enzymes is to oxidize a variety of substrates. Peroxidase exists as soluble and bound forms to the cell wall and these have been implicated in several primary and secondary metabolic processes such as phenol oxidation, chlorophyll degradation and senescence, hormone catabolism, auxin metabolism, cross linking cell wall structural proteins and polysaccharides, scavenging active oxygen radical, indoleacetic acid degradation during maturation and senescence of fruits and vegetables, lignin biosynthesis, defense mechanisms and wound healing (Brooks, 1986;Castillo et al, 1984;Gasper, 1986;Gross, 1980;Hammerchmidt et al, 1982;Pradeepkumar and Padmaja, 2008;Robinson, 1991;Scialabba et al, 2002;Thongsook and Barrett, 2005). Oxidative stress induced by chilling stress may play a pivotal role for chilling injury in plant cells (Burdon et al, 1994;Levitt, 1980;O'Kane et al, 1996;Quinn, 1988).…”

Purification and Characterization of Peroxidase from Sweet Gourd (Cucurbita moschataLam. Poiret)

“…[9,10] In addition, it is involved in plant hormone regulation, [11] defence indoleacetic acid degradation during maturation, and senescence of fruits and vegetables. [12] POD is also widely used as an important reagent for clinical diagnosis and microanalytical immunoassay. Some applications for POD have been suggested in the medicinal, chemical, and food industries.…”