Peanut Peroxidase Research Articles

The heme moiety in peanut peroxidase

Heme is present in an equimolar ratio to the apoprotein in the major cationic fraction of peanut peroxidase. The removal of heme from the holoenzyme does not affect the physicochemical and immunological properties of the apoperoxidase, however peroxidase activity is completely lost. The indoleacetic acid (IAA) oxidase activity of the apoperoxidase is reduced to 1/20 of the original holoenzyme. Both the peroxidase and IAA-oxidase activity could partially be restored in the holoenzyme reconstituted with hemin. It is suggested that heme may also participate in the IAA-oxidase activity possibly by altering the active site.

The Growth of and Peroxidase Synthesis by Two Carrot Cell Lines

The growth and the synthesis of peroxidase by two carrot lines were examined. The wild cell line WC grows more rapidly than the methotrexate resistant WCA1. However, the amount of protein released into the culture medium is identical for both cell lines while there is an approximately 10-fold greater release of peroxidase by WCA1. Since the antibodies generated against peanut peroxidase apparently react with peroxidase from carrot cells, the specific peroxidase synthesis could be determined. The ratio of newly synthesized protein to newly synthesized peroxidase confirmed the differential release of peroxidase. The results substantiate the view that there is an inverse relationship between growth and peroxidase activity of plant cells.

Immunoaffinity Studies on Cationic Peanut Peroxidase Fraction

Cationic peroxidase immobilized on sepharose has been used to separate specific antibodies from the antiperoxidase whole rabbit serum immunoglobulins (IgG) raised against it. The IgGs so separated completely pellet the peroxidase activity of the cationic peroxidase at a ratio of 6 : 1 (w/w). This shows the specificity of purified IgGs which is also shown by the single reaction arc obtained by immunoelectrophoresis of the isolated IgGs followed by challenging with crude peroxidase. These specific IgGs when linked to cyanogen bromide activated sepharose have been used to purify the cationic peroxidase from the spent medium proteins and from the cell extract of the peanut cells grown in suspension culture.

Immunological Relatedness of Peanut Peroxidase from Cells in Suspension Cultures to Peanut Plant

Summary The major cationic peroxidase released by cultured peanut cells ( Arachis hypogaea L.) and peroxidase in the peanut plant remain immunologically related. The detection of peroxidase activity in the intercellular spaces of leaves at a level eight fold higher than in leaf cells, shows its selective release in peanut plant. However, the synthesis of major cationic peroxidase is ten fold greater in the cultured cells than in the leaves.

Purification of cationic peanut peroxidase by immunoaffinity chromatography

Antibodies, raised against the major cationic peroxidase from cultured peanut cells, were coupled to CNBr-activated Sepharose to serve as the affinity source. The conditions for purification of peanut peroxidase on this antibody-coupled column were studied. The amount of peroxidase that could be purified on such affinity column depends on the purity and concentration of the antibodies coupled to Sepharose. Peroxidase from horseradish or the fungus ( Trametes versicolor) did not bind to this column. This means that the peroxidase from these sources were not immunologically identical to that of cultured peanut cells.

Some physico‐chemical properties of a major cationic peroxidase from cultured peanut cells

A major cationic peroxidase had been isolated by CMC chromatography from protein isolate of suspension medium that had supported growth of cultured peanut cells. This major cationic peroxidase proved to be antigenically different from both the anionic and the minor cationic peroxidase. Affinity for Concanavalin A found earler for the anionic peroxidase could not be detected for the major cationic peroxidase. The carbohydrate content of the major cationic peroxidase is nearly 15%. The molecular mass of the overall molecule is close to 40,000. Amino acid analysis of the hydrolysate of this major peroxidase showed similarities to amino acids of the hydrolysates of the cationic horseradish peroxidases, but no immunological relatedness could be detected between the major peanut peroxidase and the horseradish peroxidase.

Peanut and horse radish peroxidase isoenzymes. Intraspecies and interspecies immunological relatedness

AbstractThe presence of structural similarities in peroxidase isoenzymes was demonstrated by the technique of crossed immunoelec‐trophoresis. Isoenzymes found in the medium of peanut (Arachis hypogaea L.) cells grown in suspension culture were immuno‐precipitated in different arc systems according to the extent of relatedness of their antigenic determinants. Cross reactivity between isoenzymes in the medium was confirmed by demonstration that antibodies raised against only the glycoprotein fraction of the medium peroxidases would react also with the non‐glycosylated peroxidase isoenzymes that did not have affinity for Concanavalin A. Different peroxidase isoenzsmes from peanut leaves, peanut calli and peanut suspension cell medium share common antigenic determinants. Antibodies against peroxidase antigens from a different species, horse radish (Armoracia lapathifolia Gilib.), reacted with peroxidase antigens in the peanut cell medium. The results suggest a similarity of peanut peroxidases to horse radish peroxidase and. therefore, an evolutionary conservatism in peroxidase structure. The results also point to the potential utility of immunochemistry, including microcomplement fixation, to quantitate the structural similarities of peroxidase isoenzymes.

Spectral and molecular properties of peanut peroxidase isozymes

Properties of four peroxidase isozymes derived from peanut cells were examined. Electrophoresis on various concentrations of polyacrylamide gel indicated that they had the same molecular size. Filtration on Sephadex G-200 gels indicated the same Stoke's radius for all 4 isozymes. They had the same spectral properties in the oxidized, reduced and CO-reduced the pyridine hemochromogen forms, but they differed with regard to heat stability at 50° and 70° and their substrate specificity.

Iaa oxidase and polyphenol oxidase activities of peanut peroxidase isozymes

Four anionic isozymes (A 1, A 2, A 4 and A 5) from peanut cells in suspension medium possessed IAA oxidase and polyphenol oxidase activities. The specific activities of each of the enzymes differed among the 4 isozymes. The pH optima established in these assays for peroxidase was acidic, for IAA oxidase neutral and for polyphenol oxidase alkaline. All 4 isozymes had different K m and V max for the enzyme activities of peroxidase and polyphenol oxidase. The sigmoid kinetics from the IAA oxidase assays for the isozymes probably indicates an allosteric nature.