Raw reducing sugar quantification

The enzyme-enriched supernatant of P. pastoris expression was incubated with CMC solution and the release of reducing sugar by the hydrolytic process was quantified in the DNS assay. The supernatant was incubated with the cellulosic substrate for up to 20 minutes. Uninoculated YPD medium and EV-transfected inoculated medium were also detected to discard residual reducing sugars present in the reaction medium and in the basal activity of other <i>P. pastoris</i> hydrolases. TlicBgluc1 activity can enrich the reaction medium in glucose equivalents approximately twice as much as the control (2.13 mM glucose to 4.76 mM glucose).

将经毕赤酵母（P. pastoris）表达得到的酶富集上清液与羧甲基纤维素（CMC）溶液共孵育，通过DNS比色法定量检测水解过程中释放的还原糖含量。将该上清液与纤维素底物进行孵育，孵育时长最长可达20分钟。同时设置未接种的YPD培养基以及转染空载体（EV）的接种培养基作为对照，以排除反应体系中残留的还原糖以及毕赤酵母（P. pastoris）其他水解酶的本底活性对实验结果的干扰。TlicBgluc1的酶活可使反应体系中的葡萄糖当量含量约为对照组的两倍（实验组为4.76 mM葡萄糖，对照组为2.13 mM葡萄糖）。