Proenzyme Structure and Activation of Astacin Metallopeptidase
Proenzyme Structure and Activation of Astacin Metallopeptidase
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DOI：
10.1074/jbc.m109.097436
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发表时间：
2010-04-30
影响因子：
4.8
通讯作者：
Xavier Gomis-Rueth, F.
中科院分区：
文献类型：
--
作者：
Guevara, Tibisay;Yiallouros, Irene;Xavier Gomis-Rueth, F.
蛋白水解作用受无活性（潜在）酶原的调节，前段阻止底物进入酶的活性位点裂口。如何维持潜伏期通常取决于蛋白酶的催化机制。例如，在美新星金属肽酶的几个家族中，“半胱氨酸开关”机制涉及具有协调催化锌离子的半胱氨酸残基的保守前片段基序。另一个Metzincins家族，虾红素，不具有半胱氨酸开关，因此通过其他方式维持潜伏期。我们解析了欧洲小龙虾（Astacus astacus）中前列腺素的高分辨率晶体结构。其前片段是目前报道的金属肽酶中结构最短的，并且具有独特的结构。它以与真实基材相反的方向穿过活性位点裂缝。此外，由前链段的宽环投射的保守天冬氨酸协调锌离子而不是成熟酶中发现的催化溶剂分子。激活通过两步有限蛋白水解发生，并且需要柔性激活结构域的主要重排，该结构域变得刚性并形成底物结合裂口的基础。成熟还需要精确修剪新形成的 N 末端，使其能够参与埋藏的溶剂介导的氢键网络，其中包括不变的活性位点残基。我们描述了一种新的潜伏和激活机制，它与其他金属肽酶和丝氨酸肽酶具有一些共同的特征。
Proteolysis is regulated by inactive (latent) zymogens, with a prosegment preventing access of substrates to the active-site cleft of the enzyme. How latency is maintained often depends on the catalytic mechanism of the protease. For example, in several families of the metzincin metallopeptidases, a "cysteine switch" mechanism involves a conserved prosegment motif with a cysteine residue that coordinates the catalytic zinc ion. Another family of metzincins, the astacins, do not possess a cysteine switch, so latency is maintained by other means. We have solved the high resolution crystal structure of proastacin from the European crayfish, Astacus astacus. Its prosegment is the shortest structurally reported for a metallopeptidase, and it has a unique structure. It runs through the active-site cleft in reverse orientation to a genuine substrate. Moreover, a conserved aspartate, projected by a wide loop of the prosegment, coordinates the zinc ion instead of the catalytic solvent molecule found in the mature enzyme. Activation occurs through two-step limited proteolysis and entails major rearrangement of a flexible activation domain, which becomes rigid and creates the base of the substrate-binding cleft. Maturation also requires the newly formed N terminus to be precisely trimmed so that it can participate in a buried solvent-mediated hydrogen-bonding network, which includes an invariant active-site residue. We describe a novel mechanism for latency and activation, which shares some common features both with other metallopeptidases and with serine peptidases.