Structural Characterization of Arginine Fingers: Identification of an Arginine Finger for the Pyrophosphatase dUTPases.
Structural Characterization of Arginine Fingers: Identification of an Arginine Finger for the Pyrophosphatase dUTPases.
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DOI：
10.1021/jacs.6b09012
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发表时间：
2016-11
影响因子：
15
通讯作者：
Gergely N Nagy;R. Suardíaz;A. Lopata;Olivér Ozohanics;K. Vékey;B. Brooks;I. Leveles;J. Tóth;B. Vértessy;E. Rosta
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文献类型：
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作者：
Gergely N Nagy;R. Suardíaz;A. Lopata;Olivér Ozohanics;K. Vékey;B. Brooks;I. Leveles;J. Tóth;B. Vértessy;E. Rosta
精氨酸指是许多GTP酶和AAA+ATPase酶中高度保守和必需的残基，它从不同的原基完成活性部位，与核苷酸的γ-磷酸形成联系。到目前为止，还没有鉴定出使用精氨酸手指来实现上述所有性质的焦磷酸酶；所有必需的精氨酸手指都被用来催化γ-磷酸的裂解。在这里，我们识别并揭示了三聚体dUTP酶中一个保守的精氨酸残基的作用，该dUTP酶符合为精氨酸手指建立的所有标准。我们发现，与P-环状基序相邻的保守的精氨酸通过其核苷酸配位使活性中心能够进行结构组织，从而实现有效的催化，而它在过渡态稳定中的直接静电作用是次要的。对来自核苷酸水解酶和转移酶的相似的、保守的精氨酸进行了详尽的结构比较，揭示了与底物核苷酸的γ-磷酸接触的一致的氨基酸位置和方向。尽管在结构上位置相同，但dUTP酶和NTPase的精氨酸指之间的功能差异是基于焦磷酸酶dUTP酶执行的独特化学基础上的。
Arginine finger is a highly conserved and essential residue in many GTPase and AAA+ ATPase enzymes that completes the active site from a distinct protomer, forming contacts with the γ-phosphate of the nucleotide. To date, no pyrophosphatase has been identified that employs an arginine finger fulfilling all of the above properties; all essential arginine fingers are used to catalyze the cleavage of the γ-phosphate. Here, we identify and unveil the role of a conserved arginine residue in trimeric dUTPases that meets all the criteria established for arginine fingers. We found that the conserved arginine adjacent to the P-loop-like motif enables structural organization of the active site for efficient catalysis via its nucleotide coordination, while its direct electrostatic role in transition state stabilization is secondary. An exhaustive structure-based comparison of analogous, conserved arginines from nucleotide hydrolases and transferases revealed a consensus amino acid location and orientation for contacting the γ-phosphate of the substrate nucleotide. Despite the structurally equivalent position, functional differences between arginine fingers of dUTPases and NTPases are explained on the basis of the unique chemistry performed by the pyrophosphatase dUTPases.