Crystal structure of mature myroilysin and implication for its activation mechanism
Crystal structure of mature myroilysin and implication for its activation mechanism
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成熟髓溶素的晶体结构及其激活机制的意义
DOI：
10.1016/j.ijbiomac.2019.11.205
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发表时间：
2020-08-01
影响因子：
8.2
通讯作者：
Wang，Weiwu
中科院分区：
文献类型：
--
作者：
Ran，Tingting;Li，Weidong;Wang，Weiwu
Myroilysin是M12 A金属蛋白酶家族的一个新成员，具有独特的“帽”结构和独特的“半胱氨酸开关”激活机制。然而，myroilysin原的活化是难以捉摸的。在此，通过用胰蛋白酶处理myroilysin原，获得成熟myroilysin用于结构测定。成熟myroilysin的结构表明，活性位点的锌离子的成熟蛋白是协调的三个组氨酸残基，水分子，和酪氨酸残基（Tyr 208）在保守的Met-转弯基序（SIMHY）。“帽”结构从活性位点移开，使活性裂缝打开;新形成的N-末端深深地埋在myroilysin中，Glu 151直接与第一个氨基酸残基（Gly 38）形成盐桥，而它们在pro-myroilysin中彼此远离。Tyr 208的突变表明Tyr 208在myroilysin的活性中起重要作用。突变体E151 A的蛋白水解活性和热稳定性显著下降，这意味着Glu 151不仅是重要的催化，但也是至关重要的myroilysin的结构稳定性。结构比较也显示myroilysin和astacin之间存在差异。我们的生化和结构数据提供了新的见解myroilysin的激活和功能参与的关键残基Tyr 208和Glu 151。
Myroilysin is a novel bacterial member of M12A metalloproteases family with an uncommon “cysteine switch” activation mechanism and a unique “cap” structure. However, activation of pro-myroilysin is elusive. Here, mature myroilysin was obtained for structure determination by treating pro-myroilysin with trypsin. The structure of mature myroilysin showed that the active-site zinc ion of the mature protein is coordinated by three histidine residues, a water molecule, and a tyrosine residue (Tyr208) in the conserved Met-turn motif (SIMHY). The “cap” structure moves away from the active-site to leave the active cleft open; the newly formed N-terminus is deeply buried in myroilysin, and Glu151 forms a salt bridge directly with the first amino acid residue (Gly38), whereas they are far from each other in the pro-myroilysin. The mutation of Tyr208 indicates that Tyr208 plays an important role in activity of myroilysin. The proteolytic activity and thermostability of mutant E151A decreased dramatically, implying that Glu151 is not only important for catalysis, but also crucial for structural stability in myroilysin. Structural comparison also reveals differences existed between myroilysin and astacin. Our biochemical and structural data provide new insights into the activation of myroilysin and functional involvement of crucial residues Tyr208 and Glu151.