Crystal structure of the catalytic fragment of human brain 2',3'-cyclic-nucleotide 3'-phosphodiesterase.
Crystal structure of the catalytic fragment of human brain 2',3'-cyclic-nucleotide 3'-phosphodiesterase.
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DOI：
10.1016/j.jmb.2004.12.024
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发表时间：
2005-02
影响因子：
5.6
通讯作者：
Y. Sakamoto;N. Tanaka;T. Ichimiya;T. Kurihara;Kazuo T. Nakamura
中科院分区：
文献类型：
--
作者：
Y. Sakamoto;N. Tanaka;T. Ichimiya;T. Kurihara;Kazuo T. Nakamura
2′，3 ′-环核苷酸3′-磷酸二酯酶（CNP）是2 H磷酸酯酶超家族的成员，主要存在于脊椎动物的中枢神经系统，与脑白色物质紧密结合，催化2′，3 ′-环核苷酸水解生成2′-核苷酸。最近对CNP敲除小鼠的研究表明，CNP的缺失导致轴突肿胀和神经元变性。在此，以1.8 μ m分辨率解析了人CNP（hCNP-CF）的催化片段（CF）的晶体结构。它是由三个α螺旋和九个β链组成的α+β型结构。该分子的结构核心由两个拓扑等价的三链反平行β折叠组成，它们通过假2重对称性相关。每个β折叠包含H-X-T-X基序，其在2 H磷酸酯酶超家族的成员中是严格保守的。磷酸根离子与来自两个基序中的每一个的His和Thr的侧链结合。hCNP-CF与植物1″，2 ″-环核苷酸磷酸二酯酶（CPDase）和细菌2′-5′ RNA连接酶的结构比较表明，H-X-T-X基序在这些酶中是结构保守的，但活性位点的表面性质在这些酶中有很大差异，反映了它们底物的差异。根据目前的晶体结构的hCNP-CF/磷酸盐复合物，CPDase/环状核苷酸类似物复合物的可用结构，以及最近的大鼠CNP-CF的功能研究，我们提出了一种可能的底物结合模式和CNP的催化机制，它采用亲核水分子激活的His 310。所提出的反应机理基本上等同于公认的RNase A反应机理的第二步。由于hCNP-CF的整体结构与RNA酶A的结构有很大不同，因此这些酶中具有两个催化组氨酸残基的相似活性位点可能是通过趋同进化产生的。
2′,3′-Cyclic-nucleotide 3′-phosphodiesterase (CNP), a member of the 2H phosphoesterase superfamily, is firmly bound to brain white matter and found mainly in the central nervous system of vertebrates, and it catalyzes the hydrolysis of 2′,3′-cyclic nucleotide to produce 2′-nucleotide. Recent studies on CNP-knockout mice have revealed that the absence of CNP causes axonal swelling and neuronal degeneration. Here, the crystal structure of the catalytic fragment (CF) of human CNP (hCNP-CF) is solved at 1.8Å resolution. It is an α+β type structure consisting of three α-helices and nine β-strands. The structural core of the molecule is comprised of two topologically equivalent three-stranded antiparallel β-sheets that are related by a pseudo 2-fold symmetry. Each β-sheet contains an H-X-T-X motif, which is strictly conserved among members of the 2H phosphoesterase superfamily. The phosphate ion is bound to the side-chains of His and Thr from each of the two motifs. Structural comparison of hCNP-CF with plant 1″,2″-cyclic nucleotide phosphodiesterase (CPDase) and bacterial 2′-5′ RNA ligase reveals that the H-X-T-X motifs are structurally conserved among these enzymes, but the surface properties of the active site are quite different among the enzymes, reflecting the differences in their substrates. On the basis of the present crystal structure of the hCNP-CF/phosphate complex, the available structure of the CPDase/cyclic-nucleotide analogue complex, and the recent functional studies of rat CNP-CF, we propose a possible substrate-binding mode and catalytic mechanism of CNP, which employs the nucleophilic water molecule activated by His310. The proposed mechanism is basically equivalent to the second step of the well-accepted reaction mechanism of RNase A. Since the overall structure of hCNP-CF differs considerably from that of RNase A, it is likely that the similar active sites with two catalytic histidine residues in these enzymes arose through convergent evolution.