Tsai Chien-Tai, Meyer Anne S
Center for Bioprocess Engineering, Department of Chemical and Biochemical Engineering, Technical University of Denmark, DK-2800 Kgs. Lyngby, Denmark.
Molecules. 2014 Nov 25;19(12):19390-406. doi: 10.3390/molecules191219390.
The high cellulase enzyme dosages required for hydrolysis of cellulose is a major cost challenge in lignocellulosic ethanol production. One method to decrease the enzyme dosage and increase biocatalytic productivity is to re-use β-glucosidase (BG) via immobilization. In the present research, glutaraldehyde cross-linked BG was entrapped in calcium alginate gel particles. More than 60% of the enzyme activity could be recovered under optimized conditions, and glutaraldehyde cross-linking decreased leakage of BG from the calcium alginate particles. The immobilized BG aggregates were visualized by confocal laser scanning microscopy (CLSM). The CLSM images, which we believe are the first to be published, corroborate that more BG aggregates were entrapped in the matrix when the enzymes were cross-linked by glutaraldehyde as opposed to when they are not cross-linked. The particles with the immobilized BG were recycled for cellulase catalyzed hydrolysis of Avicel. No significant loss in BG activity was observed for up to 20 rounds of reaction recycle steps of the BG particles of 48 h each, verifying a significant stabilization of the BG by immobilization. Similar high glucose yields were obtained by one round of enzymatic hydrolysis of hydrothermally pretreated barley straw during a 72 h reaction with immobilized BG and free BG.
纤维素水解所需的高纤维素酶剂量是木质纤维素乙醇生产中的一个主要成本挑战。降低酶剂量并提高生物催化生产率的一种方法是通过固定化重复使用β-葡萄糖苷酶（BG）。在本研究中，戊二醛交联的BG被包埋在海藻酸钙凝胶颗粒中。在优化条件下，超过60%的酶活性可以恢复，并且戊二醛交联减少了BG从海藻酸钙颗粒中的泄漏。通过共聚焦激光扫描显微镜（CLSM）观察固定化的BG聚集体。我们认为这些CLSM图像是首次发表，证实了与未交联时相比，当酶通过戊二醛交联时，更多的BG聚集体被包埋在基质中。固定化BG的颗粒被循环用于纤维素酶催化的微晶纤维素水解。对于每轮48小时的BG颗粒反应循环步骤，高达20轮时未观察到BG活性有显著损失，证实了通过固定化BG有显著的稳定性。在72小时的反应中，使用固定化BG和游离BG对水热预处理的大麦秸秆进行一轮酶水解，获得了相似的高葡萄糖产率。
