Endoglucanase Activity Assay Protocol

Unless otherwise specified, endoglucanase activity (CMCase activity) was measured at 50°C using 1% (w/v) sodium carboxymethyl cellulose as previously described [28 (link)]. One unit of enzyme activity was defined as the amount of enzyme required to liberate 1 μM of glucose equivalent per minute under the standard assay conditions. For TaEG, the optimum temperature was determined by measuring CMCase activities at different temperatures, and thermostability was studied by incubating the enzyme at different temperatures for 1 h and then measuring the CMCase activity at 50°C. Protein concentration was determined by using a Bradford reagent kit (Sangon, China). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was performed using 12.5% (w/v) polyacrylamide gels, and proteins were stained with Coomassie Brilliant Blue R-250 (Sangon).