Structural and mutational characterization of a malate racemase from the LarA superfamily
Structural and mutational characterization of a malate racemase from the LarA superfamily
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LarA 超家族苹果酸消旋酶的结构和突变特征
DOI：
10.1007/s10534-022-00372-x
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发表时间：
2022
期刊：
影响因子：
3.5
通讯作者：
Hu, Jian
中科院分区：
文献类型：
--
作者：
Gatreddi, Santhosh;Urdiain-Arraiza, Julian;Desguin, Benoît;Hausinger, Robert P.;Hu, Jian
LarA超家族由使用多种α-羟基酸催化外消旋/差向异构反应的镍依赖性酶组成。首先表征的LarA是一种来自植物乳杆菌的乳酸消旋酶，导致发现了镍钳核苷酸（NPN）辅因子，该辅因子被家族成员利用替代底物，包括来自嗜热厌氧杆菌（Thermoanaerobacterium thermosaccharolyticum）（Mar 2）的苹果酸消旋酶。在这项工作中，获得了Mar 2的更高分辨率的晶体结构和更好的数据质量，揭示了该蛋白质新的结构和动力学特征。一个结合辅因子和底物的Mar 2结构模型被生成，以揭示LarA超家族中两个不同亚群之间的共同和独特特征。此外，使用结构指导的突变研究来检查被建模为与NPN相互作用的残基的重要性，并探索哪些残基对于赋予苹果酸特异性至关重要。特别是，在Mar 2中参与底物结合的两个残基的取代以匹配LarA中的相应残基导致获得低水平的乳酸消旋酶活性。额外的兴趣，底物谱扩展到包括酒石酸盐，苹果酸盐的类似物。这些新发现将有助于更好地了解广泛分布于细菌和古细菌物种中的许多其他LarA同系物的结构-功能关系。
The LarA superfamily consists of nickel-dependent enzymes catalyzing racemization/epimerization reactions using a variety of α-hydroxy acids. The first-characterized LarA, a lactate racemase fromLactobacillus plantarum, led to the discovery of the nickel-pincer nucleotide (NPN) cofactor that is utilized by family members with alternative substrates, including malate racemase fromThermoanaerobacterium thermosaccharolyticum(Mar2). In this work, a higher resolution crystal structure of Mar2 was obtained with better data quality that revealed new structural and dynamic characteristics of the protein. A model of the Mar2 structure with bound cofactor and substrate was generated to uncover the common and the unique features among two distinct subgroups in the LarA superfamily. In addition, structure-guided mutational studies were used to examine the importance of residues that are modeled to interact with NPN and to explore which residues were critical for conferring specificity for malate. In particular, substitution of two residues involved in substrate binding in Mar2 to match the corresponding residues in LarA led to the acquisition of low levels of lactate racemase activity. Of additional interest, the substrate spectrum was expanded to include tartrate, an analog of malate. These new findings will help to better understand structure–function relationships of many other LarA homologs that are broadly distributed in bacterial and archaeal species.
DOI：
10.1107/s2059798319011471
发表时间：
2019-10-01
影响因子：
2.2
作者：
Liebschner, Dorothee;Afonine, Pavel V.;Adams, Paul D.
通讯作者：
Adams, Paul D.