Visualizing active-site dynamics in single crystals of HePTP: opening of the WPD loop involves coordinated movement of the E loop.
Visualizing active-site dynamics in single crystals of HePTP: opening of the WPD loop involves coordinated movement of the E loop.
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DOI：
10.1016/j.jmb.2010.11.020
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发表时间：
2011-01-21
影响因子：
5.6
通讯作者：
Page R
中科院分区：
文献类型：
--
作者：
Critton DA;Tautz L;Page R
蛋白酪氨酸磷酸酶 (PTP) 水解磷酸酪氨酸涉及 PTP 环的底物结合和 WPD 环对活性位点的封闭。 E 环紧邻 PTP 和 WPD 环，在人类 PTP 中的序列和结构都是保守的，但该环在底物结合/催化中的作用相对而言尚未被探索。造血酪氨酸磷酸酶 (HePTP) 是激酶相互作用基序 (KIM)-PTP 家族的成员。与其他 PTP 相比，KIM-PTP 具有序列和结构独特的 E 环。为了了解 E 环在 HePTP 闭合状态和开放状态之间转变中的作用，我们鉴定了 HePTP 的一种新晶型，它允许在单个晶型内观察到闭合状态到开放状态的转变。这些结构（包括 HePTP 开放状态的第一个结构）表明 WPD 环采用“非典型开放”构象，重要的是，配体可以在活性位点交换，这对于 HePTP 抑制剂的开发至关重要。这些结构还表明，四面体含氧阴离子结合在一个新的二级位点，并发挥协调 PTP、WPD 和 E 环的功能。最后，利用结构和动力学数据，我们揭示了 E 环残基 Lys182 通过与 WPD 环的 Asp236 相互作用在增强 HePTP 催化活性中的新作用，为催化循环中 WPD 和 E 环的协调动力学提供了第一个证据，正如我们所表明的，这与多个 PTP 家族相关。
Phosphotyrosine hydrolysis by protein tyrosine phosphatases (PTPs) involves substrate binding by the PTP loop and closure over the active site by the WPD loop. The E loop, located immediately adjacent to the PTP and WPD loops, is conserved among human PTPs in both sequence and structure, yet the role of this loop in substrate binding/catalysis is comparatively unexplored. Hematopoietic tyrosine phosphatase (HePTP) is a member of the kinase interaction motif (KIM)-PTP family. Compared to the other PTPs, the KIM-PTPs have E loops that are unique in both sequence and structure. In order to understand the role of the E loop in the transition between the closed and open states of HePTP, we identified a novel crystal form of HePTP that allowed the closed-to-open state transition to be observed within a single crystal form. These structures, which include the first structure of the HePTP open state, show that the WPD loop adopts an ‘atypically open’ conformation and, importantly, that ligands can be exchanged at the active site, critical for HePTP inhibitor development. These structures also show that tetrahedral oxyanions bind at a novel, secondary site and function to coordinate the PTP, WPD and E loops. Finally, using both structural and kinetic data, we reveal a novel role for E loop residue Lys182 in enhancing HePTP catalytic activity through its interaction with Asp236 of the WPD loop, providing the first evidence for coordinated dynamics of the WPD and E loops in the catalytic cycle which, as we show, are relevant to multiple PTP families.
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影响因子：
16
作者：
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通讯作者：
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DOI：
10.1107/s0907444904019158
发表时间：
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DOI：
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发表时间：
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期刊：
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影响因子：
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作者：
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通讯作者：
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DOI：
10.1002/prot.10286
发表时间：
2003-02-15
期刊：
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影响因子：
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通讯作者：
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