Characterization of beta-Glucosidase Isoenzymes Possibly Involved in Lignification from Chick Pea (Cicer arietinum L.) Cell Suspension Cultures

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Abstract
Crude cell wall preparations from Cicer arietinum L. cell suspension cultures show high activity for the hydrolysis of coniferyl alcohol β-d-glucoside (coniferin). Various β-glucosidase activities could be solubilized from these preparations by 0.5 M NaCl treatment and one of these could be shown to possess a high activity for the hydrolysis of coniferin. The enzyme activities were purified to near homogeneity by Sephadex G-200 and CM-Sephadex chromatography. Isoelectric focussing indicated the occurrence of β-glucosidase isoenzymes with identical catalytic activity (pI 8.5–10). Molecular weights were determined as 110 000, with two subunits of 63000 and 43000. Maximum hydrolytic activity is at pH 5. The β-glucosidase isoenzymes catalyze the hydrolysis of various β-glucosides with aromatic aglycone structure and different sugar moieties. However, coniferin has been found to be one of the best substrates (km= 0.8 mM; V= 6 μmol · min−1· mg protein−1) for these β-glucosidase isoenzymes. The data suggest that a β-glucosidase-catalyzed reaction might be involved in lignification of these plant cell cultures.