Purification And Characterization Of β-1, 3-Glucanase From Highland Barley And Preparation Of Its Antibody

The β-1,3-glucanase[EC 3.2.1.39]are abundant, highly regulated enzymes, widely distributed in seed-plant species. They are able to catalyse hydrolytic cleavage of the 1,3-β-D-glucosidic linkages in β-1,3-glucans. The β-1,3-glucanase exist as a family of multiple isoforms that differ in size , isoelectric point, primary structure, cellular localization and pattern of regulation. There is strong envidence that these enzyme are implicated in diverse physiological and developmental processes in the uninfected plant, including cell division, fertilization, embryo genesis, seed germination. β-1,3 -gluca naseare also induced in response to the infection of plants with pathogens and are therefore, grouped among the pathogenesis -related (PR) proteins as the PR-2 family.In this paper, a major isoform of β-1,3-glucanase from highland barley seedings has been purified by fractional precipitation with 50 %～85% ammonium sulfate, DEAE-Cellulose 52 and CM-Sepharose (Fast Flow) ion-exchange chromatography and Sephadex G-75. The molecular weight of β-1,3 -glucanase was estimated about 27 kDa by SDS-PAGE. β-1,3- glucanase has pH optimum of 5.5 with laminarin and was stable up to 40 ℃ .With laminarin ,the enzyme had a Km of 0.59 mg/mL and tempreture optimum of 35 ℃. When treated with Ca²⁺, Pb²⁺ at different concentrations, activity assay and fluor escence spectra show that β-1,3-glucanase...