Crystal structures of recombinant human purple acid phosphatase with and without an inhibitory conformation of the repression loop
Crystal structures of recombinant human purple acid phosphatase with and without an inhibitory conformation of the repression loop
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DOI：
10.1016/j.jmb.2005.04.014
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发表时间：
2005-08-05
影响因子：
5.6
通讯作者：
Guss, JM
中科院分区：
文献类型：
--
作者：
Sträter, N;Jasper, B;Guss, JM
在大肠杆菌和毕赤酵母中重组表达的人紫色酸性磷酸酶（rHPAPEJ和rHPAPpp）在2.2埃分辨率下以两种不同的晶体形式进行了晶体结构的测定。在这两种情况下，酶在氧化（无活性）状态下结晶，其中双核活性位点的两个铁原子都是铁（III）。这两种结构的主要区别在于酶“抑制环”的构象。该环在体内或体外的蛋白水解裂解可显著激活哺乳动物的pap。在从rHPAPE获得的晶体中，该环的Asp145的羧酸侧链作为双齿配体连接两个金属原子，其方式类似于酶-底物复合物中磷酸酯底物的可能结合模式。因此，Asp145的羧酸侧链和邻近的Phe146侧链阻断了活性位点，从而使酶失活。在rHPAP（Pp）的晶体结构中，酶11抑制环具有类似于在其他哺乳动物PAP结构中观察到的开放构象。目前的结构表明抑制环具有显著的构象灵活性；观察到的交替结合模式表明该环可能具有抑制作用。(c) 2005 Elsevier Ltd版权所有。
The crystal structure of human purple acid phosphatase recombinantly expressed in Escherichia coli (rHPAPEJ and Pichia pastoris (rHPAPpp) has been determined in two different crystal forms, both at 2.2 angstrom resolution. In both cases, the enzyme crystallized in its oxidized (inactive) state, in which both Fe atoms in the dinuclear active site are Fe(III). The main difference between the two structures is the conformation of the enzyme "repression loop". Proteolytic cleavage of this loop in vivo or in vitro results in significant activation of the mammalian PAPs. In the crystals obtained from rHPAPE, the carboxylate side-chain of Asp145 of this loop acts as a bidentate ligand that bridges the two metal atoms, in a manner analogous to a possible binding mode for a phosphate ester substrate in the enzyme-substrate complex. The carboxylate side-chain of Asp145 and the neighboring Phe146 side-chain thus block the active site, thereby inactivating the enzyme. In the crystal structure of rHPAP(Pp), the enzyme 11 repression loop" has an open conformation similar to that observed in other mammalian PAP structures. The present structures demonstrate that the repression loop exhibits significant conformational flexibility; and the observed alternate binding mode suggests a possible inhibitory role for this loop. (c) 2005 Elsevier Ltd. All rights reserved.