Structure-based design of agarase AgWH50C from Agarivorans gilvus WH0801 to enhance thermostability
Structure-based design of agarase AgWH50C from Agarivorans gilvus WH0801 to enhance thermostability
复制标题
DOI：
10.1007/s00253-018-9540-1
复制
发表时间：
2018-12
影响因子：
5
通讯作者：
Pujuan Zhang;Jinru Zhang;Lujia Zhang;Jianan Sun;Yuan Li;Lian Wu;Jiahai Zhou;C. Xue;Xiangzhao Mao-X
中科院分区：
文献类型：
--
作者：
Pujuan Zhang;Jinru Zhang;Lujia Zhang;Jianan Sun;Yuan Li;Lian Wu;Jiahai Zhou;C. Xue;Xiangzhao Mao-X
AgWH 50 C是从Agarivorans gilvusWH 0801中分离到的GH 50的外切β-琼胶酶，在新琼胶二糖的酶促生产中起着关键作用，在化妆品和医药工业中具有广阔的应用前景。相反，较差的热稳定性成为糖苷水解酶（GH）家族50种琼脂糖酶（包括AgWH 50 C）的主要阻碍因素。在此，基于AgWH 50 C的晶体结构，我们设计了几个突变体，通过多重交联的合理设计方案，使用热稳定性预测软件ETSS，PoPMuSiC，和HotMuSiC。令我们惊讶的是，突变体K621 F与野生型AgWH 50 C（30 °C）相比，其相对活性增加了45%，最适温度增加到38 °C。K621 F的热稳定性也表现出实质性的改善。考虑到琼脂糖的胶凝温度高于35 °C，K621 F可用于水解琼脂糖生产新琼脂二糖。
AgWH50C, an exo-β-agarase of GH50 isolated fromAgarivorans gilvusWH0801, plays a key role in the enzymatic production of neoagarobiose, which has great application prospect in the cosmetics and pharmaceutical industry. In contrast, the poor thermostability becomes the main obstructive factor of glycoside hydrolase (GH) family 50 agarases, including AgWH50C. Herein, based on the AgWH50C crystal structure, we designed several mutants by a multiple cross-linked rational design protocol used thermostability predicting softwares ETSS, PoPMuSiC, and HotMuSiC. To our surprise, the mutant K621F increased its relative activity by as much as 45% and the optimal temperature increased to 38 °C compared to that of wild-type, AgWH50C (30 °C). The thermostability of K621F also exhibited a substantial improvement. Considering that the gelling temperature of the agarose is higher than 35 °C, K621F can be used to hydrolyze agarose for neoagarobiose production.