Novel Antarctic Endo-Polygalacturonase for Pectin Extraction and Vegetal Tissue Maceration at Mild Temperatures.

Abstract

The aim of the present work was to partially purify and characterize an Antarctic polygalacturonase and to determine the enzyme's potential in pectin extraction and vegetal maceration at 20 °C. Polygalacturonase was purified by chromatography to obtain an enzymatic preparation of specific activity 30.3 U.mg^-1. Optimal conditions for the polygalacturonase activity were 45 °C and pH 5.0-6.0, and the activation energy for the reaction was 41.8 kJ.mol^-1. Of the enzyme activity, 100% was retained after 3 h at 40 °C. The enzyme was remarkably stable for an hour over a wide range of pH (2.0-12.0). Polygalacturonase activity was slightly reduced in the presence of Ca⁺², Fe⁺³, K⁺, Mn⁺², and Zn⁺², whereas Hg⁺² reduced the activity by 60%, suggesting a thiol-dependent catalysis. The apparent molecular weight of the enzyme was 33 kDa. The kinetic constants evaluated against polygalacturonic acid were 0.17 mg.ml^-1 (K_m), 480 s^-1 (K_cat), and 7.9 µmol.mg^-1.min^-1 (V_max). The enzyme was active against different pectic substrates. Thin-layer chromatography revealed an endo-mechanism of action. Polygalacturonase digested lime pomace to aid the extraction of high-methoxylated pectin at 20 °C and increased the vegetal maceration of Capsicum annuum by 24% over the control values.