Purification and characterization of a neutral peroxidase induced by rubbing tomato internodes

Abstract

Rubbing applied to a young tomato internode inhibited the elongation of this internode and increased soluble peroxidase activity. These morphological and biochemical changes were observed both at the site of rubbing (local response) and in the neighbouring internode (systemic response). The cellular, biochemical, and molecular mechanisms leading to inhibition of internode elongation are not fully understood. It was proposed that mechanical stimuli increased the oxidation of IAA, via the induction of specific peroxidases and stimulated the lignification processes. In order to gain more information about the role of these enzymes, analysis of changes in peroxidase activities were performed. Qualitative analysis of isoperoxidases, by means of native cathodic PAGE, showed four induced isoforms termed C1, C2, C3, and C4. The major isoform (C2) was purified to homogeneity and partially characterized. This isoform is probably unglycosylated, with a molecular mass of 36 kDa and a neutral pI of 7.1. The effects of pH and temperature on the activity were determined with guaiacol as electron donor. Optima were obtained at pH 5 and at a temperature of 55°C. The activity of the purified enzyme was not affected by Ca2+, Mg2+ and Mn2+ as was reported for some basic peroxidases. Analysis of substrate specificity revealed that this isoperoxidase acted on ABTS, o‐dianisidine, pyrogallol, guaiacol, coniferyl alcohol (monolignol) and IAA but not on syringaldazine. Activitiy of C2 isoperoxidase on coniferyl alcohol and IAA suggests a possible role of peroxidase C2 in inhibition of internode elongation, observed in rubbed plants, probably via an increase in lignification processes and regulation of IAA levels in internode tissues.