The active site of Serratia endonuclease contains a conserved magnesium-water cluster
The active site of Serratia endonuclease contains a conserved magnesium-water cluster
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DOI：
10.1006/jmbi.1999.2729
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发表时间：
1999-05-21
影响因子：
5.6
通讯作者：
Krause, KL
中科院分区：
文献类型：
--
作者：
Miller, MD;Cai, JW;Krause, KL
沙雷氏菌核酸内切酶是自然界中广泛分布的一类镁依赖性核酸酶的重要成员。在这里，我们描述了该酶活性位点内镁-水簇的位置和几何形状。镁原子的唯一蛋白质配体是 Asn119；该金属离子还与五个水分子结合以形成八面体配位络合物。这些水分子非常有序，没有旋转紊乱或运动的证据。 Glu127 和 His89 位于附近，并且各自与配位层中的水分子形成氢键。 Asp86 不与镁或其周围的水分子螯合。动力学和定点诱变实验的结果表明，该金属-水簇含有该酶的催化金属离子。与协调镁原子的水分子形成氢键的所有残基在与沙雷氏菌核酸内切酶同源的核酸酶中都是保守的，这表明水簇是该酶家族的保守特征。我们提供了与另一种核酸酶（归巢核酸内切酶 I-PpoI）的详细结构比较，最近显示，尽管缺乏序列同源性，但它与沙雷氏菌核酸内切酶具有相似的活性位点几何结构。这两种结构的证据表明沙雷氏菌核酸酶的镁通过内球机制参与催化。 (C) 1999 年学术出版社。
Serratia endonuclease is an important member of a class of magnesium dependent nucleases that are widely distributed in nature. Here, we describe the location and geometry of a magnesium-water cluster within the active site of this enzyme. The sole protein ligand of the magnesium atom is Asn119; this metal ion is also associated with five water molecules to complete an octahedral coordination complex. These water molecules are very well ordered and there is no evidence of rotational disorder or motion. Glu127 and His89 are located nearby and each is hydrogen bonded to water molecules in the coordination sphere. Asp86 is not chelated to the magnesium or its surrounding water molecules. Results of kinetics and site-specific mutagenesis experiments suggest that this metal-water cluster contains the catalytic metal ion of this enzyme. All residues which hydrogen bond to the water molecules that coordinate the magnesium atom are conserved in nucleases homologous to Serratia endonuclease, suggesting that the water cluster is a conserved feature of this family of enzymes. We offer a detailed structural comparison to one other nuclease, the homing endonuclease I-PpoI, that has recently been shown, in spite of a lack of sequence homology, to share a similar active site geometry to Serratia endonuclease. Evidence from both of these structures suggests that the magnesium of Serratia nuclease participates in catalysis via an inner sphere mechanism. (C) 1999 Academic Press.