Structural insight into the catalytic mechanism of gluconate 5-dehydrogenase from Streptococcus suis: Crystal structures of the substrate-free and quaternary complex enzymes
Structural insight into the catalytic mechanism of gluconate 5-dehydrogenase from Streptococcus suis: Crystal structures of the substrate-free and quaternary complex enzymes
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猪链球菌葡萄糖酸 5-脱氢酶催化机制的结构洞察：无底物和四元复合酶的晶体结构
DOI：
10.1002/pro.32
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发表时间：
2009-02-01
期刊：
影响因子：
8
通讯作者：
Gao, George F.
中科院分区：
文献类型：
--
作者：
Zhang, Qiangmin;Peng, Hao;Gao, George F.
葡萄糖酸5-脱氢酶（Ga 5DH）是一种NADP（H）依赖性酶，催化D-葡萄糖酸和5-酮-D-葡萄糖酸之间的可逆氧化还原反应，从而调节细菌中这种重要碳和能源的流量。尽管有相当多的生理和生化知识的Ga 5DH，有很少的物理或结构信息可用于这种酶。为此，我们在此报告的晶体结构的Ga 5DH从致病性猪链球菌血清型2在无底物和配体（NADP 1/D-葡萄糖酸盐/金属离子）四元复合物的形式在2.0埃分辨率。结构分析表明，Ga 5DH采用类似于短链脱氢酶/还原酶（SDR）家族成员中发现的蛋白质折叠，而酶本身代表了该家族以前未表征的成员。在溶液中，Ga 5DH作为四聚体存在，其包含四个相同的类似于29 kDa的亚基。Ga 5DH的催化位点显示出与SDR家族的其他酶相当大的结构相似性，但S. suis蛋白含有一个额外的残基（Arg 104），该残基在底物的结合和定位中起重要作用。四元复合物结构提供了第一个明确的晶体学证据的催化重要的丝氨酸残基的作用，也揭示了一个氨基酸四分体RSYK，不同于SYK三联体中发现的大多数SDR酶。晶体结构的详细分析揭示了重要的贡献的Ca 2+离子的活性位点的形成和特定的残基在C-末端的亚基四聚体组装。由于Ga 5DH是一个潜在的治疗靶点，我们的研究结果不仅提供了催化机制的见解，而且还提出了基于结构的药物设计的靶点。
Gluconate 5-dehydrogenase (Ga5DH) is an NADP(H)-dependent enzyme that catalyzes a reversible oxidoreduction reaction between D-gluconate and 5-keto-D-gluconate, thereby regulating the flux of this important carbon and energy source in bacteria. Despite the considerable amount of physiological and biochemical knowledge of Ga5DH, there is little physical or structural information available for this enzyme. To this end, we herein report the crystal structures of Ga5DH from pathogenic Streptococcus suis serotype 2 in both substrate-free and liganded (NADP1/D-gluconate/metalion) quaternary complex forms at 2.0 angstrom resolution. Structural analysis reveals that Ga5DH adopts a protein fold similar to that found in members of the short chain dehydrogenase/reductase (SDR) family, while the enzyme itself represents a previously uncharacterized member of this family. In solution, Ga5DH exists as a tetramer that comprised four identical similar to 29 kDa subunits. The catalytic site of Ga5DH shows considerable architectural similarity to that found in other enzymes of the SDR family, but the S. suis protein contains an additional residue (Arg104) that plays an important role in the binding and orientation of substrate. The quaternary complex structure provides the first clear crystallographic evidence for the role of a catalytically important serine residue and also reveals an amino acid tetrad RSYK that differs from the SYK triad found in the majority of SDR enzymes. Detailed analysis of the crystal structures reveals important contributions of Ca2+ ions to active site formation and of specific residues at the C-termini of subunits to tetramer assembly. Because Ga5DH is a potential target for therapy, our findings provide insight not only of catalytic mechanism, but also suggest a target of structure-based drug design.