Crystal structure of human arylsulfatase A: The aldehyde function and the metal ion at the active site suggest a novel mechanism for sulfate ester hydrolysis
Crystal structure of human arylsulfatase A: The aldehyde function and the metal ion at the active site suggest a novel mechanism for sulfate ester hydrolysis
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DOI：
10.1021/bi9714924
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发表时间：
1998-03-17
期刊：
影响因子：
2.9
通讯作者：
Saenger, W
中科院分区：
文献类型：
--
作者：
Lukatela, G;Krauss, N;Saenger, W
人溶酶体芳基硫酸酯酶 A (ASA) 是硫酸酯酶家族的原型成员，这些酶需要将保守半胱氨酸的 -CH2SH 基团翻译后氧化为醛，产生甲酰甘氨酸。如果没有这种修饰，硫酸酯酶就没有催化活性，如被称为多种硫酸酯酶缺乏症的溶酶体贮积症所揭示的那样。 2.1 埃分辨率的 X 射线晶体结构显示由二聚体 (α(2))(4) 四聚体组成的 ASA 同八聚体。该单体的α/β折叠与另一种水解酶碱性磷酸酶具有显着的结构相似性，并且这两种结构的叠加表明活性中心位于大致相同的位置。功能必需的甲酰甘氨酸位于带正电荷的口袋中，并充当八面体配位金属离子（解释为 Mg2+）的配体。甲酰甘氨酸处的电子密度表明存在 2 倍无序醛基，其中可能含有带有宝石羟基的醛水合物 -CH(OH)(2)。在所提出的催化机制中，醛接受水分子形成水合物。两个羟基之一通过酯交换步骤水解底物硫酸酯，产生共价中间体。第二个羟基用于通过C-O裂解和醛的重组在构型反转下消除硫酸盐。这项研究为理解酯水解的新机制提供了结构基础，并解释了异常修饰氨基酸的功能重要性。
Human lysosomal arylsulfatase A (ASA) is a prototype member of the sulfatase family, These enzymes require the posttranslational oxidation of the -CH2SH group of a conserved cysteine to an aldehyde, yielding a formylglycine. Without this modification sulfatases are catalytically inactive, as revealed by a lysosomal storage disorder known as multiple sulfatase deficiency. The 2.1 Angstrom resolution X-ray crystal structure shows an ASA homooctamer composed of a tetramer of dimers, (alpha(2))(4). The alpha/beta fold of the monomer has significant structural analogy to another hydrolytic enzyme, the alkaline phosphatase, and superposition of these two structures shows that the active centers are located in largely identical positions. The functionally essential formylglycine is located in a positively charged pocket and acts as ligand to an octahedrally coordinated metal ion interpreted as Mg2+. The electron density at the formylglycine suggests the presence of a 2-fold disordered aldehyde group with the possible contribution of an aldehyde hydrate, -CH(OH)(2), with gem-hydroxyl groups. In the proposed catalytic mechanism, the aldehyde accepts a water molecule to form a hydrate. One of the two hydroxyl groups hydrolyzes the substrate sulfate ester via a transesterification step, resulting in a covalent intermediate, The second hydroxyl serves to eliminate sulfate under inversion of configuration through C-O cleavage and reformation of the aldehyde. This study provides the structural basis for understanding a novel mechanism of ester hydrolysis and explains the functional importance of the unusually modified amino acid.