Zhou Peng, Chen Zhongzhou, Yan Qiaojuan, Yang Shaoqing, Hilgenfeld Rolf, Jiang Zhengqiang
Department of Biotechnology, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, People's Republic of China.
Acta Crystallogr D Biol Crystallogr. 2013 Oct;69(Pt 10):2027-38. doi: 10.1107/S090744491301799X. Epub 2013 Sep 20.
Endo-β-1,3-glucanases catalyze the hydrolysis of β-1,3-glycosidic linkages in glucans. They are also responsible for rather diverse physiological functions such as carbon utilization, cell-wall organization and pathogen defence. Glycoside hydrolase (GH) family 81 mainly consists of β-1,3-glucanases from fungi, higher plants and bacteria. A novel GH family 81 β-1,3-glucanase gene (RmLam81A) from Rhizomucor miehei was expressed in Escherichia coli. Purified RmLam81A was crystallized and the structure was determined in two crystal forms (form I-free and form II-Se) at 2.3 and 2.0 Å resolution, respectively. Here, the crystal structure of a member of GH family 81 is reported for the first time. The structure of RmLam81A is greatly different from all endo-β-1,3-glucanase structures available in the Protein Data Bank. The overall structure of the RmLam81A monomer consists of an N-terminal β-sandwich domain, a C-terminal (α/α)6 domain and an additional domain between them. Glu553 and Glu557 are proposed to serve as the proton donor and basic catalyst, respectively, in a single-displacement mechanism. In addition, Tyr386, Tyr482 and Ser554 possibly contribute to both the position or the ionization state of the basic catalyst Glu557. The first crystal structure of a GH family 81 member will be helpful in the study of the GH family 81 proteins and endo-β-1,3-glucanases.
内切β-1,3-葡聚糖酶催化葡聚糖中β-1,3-糖苷键的水解。它们还负责多种生理功能，如碳利用、细胞壁组织和病原体防御。糖苷水解酶（GH）家族81主要由来自真菌、高等植物和细菌的β-1,3-葡聚糖酶组成。来自米黑根毛霉的一个新的GH家族81β-1,3-葡聚糖酶基因（RmLam81A）在大肠杆菌中表达。纯化后的RmLam81A被结晶，并分别以2.3 Å和2.0 Å的分辨率确定了两种晶体形式（形式I-游离和形式II-硒）的结构。在此，首次报道了GH家族81成员的晶体结构。RmLam81A的结构与蛋白质数据库中所有可用的内切β-1,3-葡聚糖酶结构有很大不同。RmLam81A单体的整体结构由一个N端β-折叠结构域、一个C端（α/α）6结构域以及它们之间的一个额外结构域组成。在单置换机制中，Glu553和Glu557分别被认为是质子供体和碱性催化剂。此外，Tyr386、Tyr482和Ser554可能对碱性催化剂Glu557的位置或电离状态都有贡献。GH家族81成员的首个晶体结构将有助于对GH家族81蛋白质和内切β-1,3-葡聚糖酶的研究。
