Preparation and purification of glucanase and chitinase from bean leaves

Glucanase (endo-beta-1, 3-glucan 3-glucanohydrolase, EC 3.2.1.6, laminarinase, callase) and chitinase (poly-beta-1, 4-[2-acetamido-2-deoxy] -d-glucoside glycanohydrolase, EC 3.2.1.14) were extracted from ethylene-treated bean (Phaseolus vulgaris L. cv. Red Kidney) leaves and purified on hydroxyapatite and carboxymethyl Sephadex columns. The glucanase prepared was homogeneous as judged by analytical centrifugation data, electrophoresis, and antibody-antigen reactions. On the basis of gel filtration, antibody-antigen reactions, and amino acid analysis, the molecular weight was estimated to be between 11,500 and 12,500. However, ultracentrifugation gave a higher estimate of 34,000. The glucanase had an isoelectric point near pH 11 and was specific for beta-1, 3-linkages. The chitinase was only partially purified as judged by electrophoretic behavior.