Gong W, Zhu X, Liu S, Teng M, Niu L
Department of Molecular Biology and Cell Biology and Laboratory of Structural Biology, School of Life Science, University of Science and Technology of China, Hefei, Anhui, 230026, P.R. China.
J Mol Biol. 1998 Oct 30;283(3):657-68. doi: 10.1006/jmbi.1998.2110.
Acutolysin A alias AaHI, a 22 kDa hemorrhagic toxin isolated from the snake venom of Agkistrodon acutus, is a member of the adamalysin subfamily of the metzincin family and is a snake venom zinc metalloproteinase possessing only one catalytic domain. Acutolysin A was found to have a high-activity and a low-activity under weakly alkaline and acidic conditions, respectively. With the adamalysin II structure as the initial trial-and-error model, the crystal structures were solved to the final crystallographic R-factors of 0. 168 and 0.171, against the diffraction data of crystals grown under pH 5.0 and pH 7.5 conditions to 1.9 A and 1.95 A resolution, respectively. One zinc ion, binding in the active-site, one structural calcium ion and some water molecules were localized in both of the structures. The catalytic zinc ion is coordinated in a tetrahedral manner with one catalytic water molecule anchoring to an intermediate glutamic acid residue (Glu143) and three imidazole Nepsilon2 atoms of His142, His146 and His152 in the highly conserved sequence H142E143XXH146XXGXXH152. There are two new disulfide bridges (Cys157-Cys181 and Cys159-Cys164) in acutolysin A in addition to the highly conserved disulfide bridge Cys117-Cys197. The calcium ion occurs on the molecular surface. The superposition showed that there was no significant conformational changes between the two structures except for a few slight changes of some flexible residue side-chains on the molecular surface, terminal residues and the active-site cleft. The average contact distance between the catalytic water molecule and oxygen atoms of the Glu143 carboxylate group in the weakly alkaline structure was also found to be closer than that in the weakly acidic structure. By comparing the available structural information of the members of the adamalysin subfamily, it seems that, when lowering the pH value, the polarization capability of the Glu143 carboxylate group to the catalytic water molecule become weaker, which might be the structural reason why the snake venom metalloproteinases are inactive or have a low activity under acidic conditions.
尖吻蝮溶血素A别名AaHI，是一种从尖吻蝮蛇毒中分离出的22 kDa出血毒素，是金属锌蛋白酶家族中解整合素样金属蛋白酶亚家族的成员，是一种仅具有一个催化结构域的蛇毒锌金属蛋白酶。已发现尖吻蝮溶血素A在弱碱性和酸性条件下分别具有高活性和低活性。以解整合素II结构作为初始试错模型，针对在pH 5.0和pH 7.5条件下生长的晶体的衍射数据，分别以1.9 Å和1.95 Å分辨率解析了晶体结构，最终晶体学R因子分别为0.168和0.171。在这两种结构中均定位了一个结合在活性位点的锌离子、一个结构钙离子和一些水分子。催化锌离子以四面体方式与一个催化水分子配位，该水分子锚定在高度保守序列H142E143XXH146XXGXXH152中的中间谷氨酸残基（Glu143）以及His142、His146和His152的三个咪唑Nε2原子上。除了高度保守的二硫键Cys117-Cys197外，尖吻蝮溶血素A中还有两个新的二硫键（Cys157-Cys181和Cys159-Cys164）。钙离子位于分子表面。叠加显示，除了分子表面上一些柔性残基侧链、末端残基和活性位点裂隙有一些轻微变化外，两种结构之间没有明显的构象变化。还发现弱碱性结构中催化水分子与Glu143羧酸盐基团的氧原子之间的平均接触距离比弱酸性结构中的更近。通过比较解整合素亚家族成员的现有结构信息，似乎在降低pH值时，Glu143羧酸盐基团对催化水分子的极化能力变弱，这可能是蛇毒金属蛋白酶在酸性条件下无活性或活性低的结构原因。
