Quaternary structure of α-amino-β-carboxymuconate-E-semialdehyde decarboxylase (ACMSD) controls its activity
Quaternary structure of α-amino-β-carboxymuconate-E-semialdehyde decarboxylase (ACMSD) controls its activity
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DOI：
10.1074/jbc.ra119.009035
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发表时间：
2019-07-26
影响因子：
4.8
通讯作者：
Liu, Aimin
中科院分区：
文献类型：
--
作者：
Yang, Yu;Davis, Ian;Liu, Aimin
Alpha-Amino-beta-carboxymuconate-E-semialdehyde脱羧酶通过犬尿氨酸途径在L色氨酸降解过程中起重要作用。ACMSD形成同源二聚体，作为单体在功能上是不活跃的，因为它的催化组装需要邻近亚基的精氨酸残基。然而，ACMSD在溶液中如何控制低聚状态和自缔合仍未被探索。在这里，我们证明了来自荧光假单胞菌的ACMSD可以自组装成同源二聚体、四聚体和更高阶结构。利用尺寸排斥色谱结合小角X射线散射(SEC-SAXS)分析，研究了ACMSD四聚体的结构，并将SAXS数据与单体组分的X射线晶体结构进行了拟合，得到了该四聚体的准原子结构。这一分析揭示了ACMSD的四聚体模型，即二聚体的正面二聚体。我们观察到四聚体的催化活性比二聚体更高，并且与单体和二聚体处于平衡状态。取代二聚体-二聚体界面的关键残基His-110，在不改变X-射线晶体结构的情况下，通过增加溶液中高阶齐聚物的比例改变了四聚体的解离轮廓。ACMSD的自缔合受到pH、离子强度和其他静电相互作用的影响。对ACMSD序列的比对表明，His-110在少数利用硝基苯甲酸作为唯一碳和能量来源的细菌中高度保守，这表明ACMSD在四聚体和高阶结构中的自组装具有专门的功能作用。这些结果表明，动态齐聚状态可能调节ACMSD的活性，SEC-SAXS与X射线结晶学相结合是研究蛋白质自结合的有力工具。
alpha-Amino-beta-carboxymuconate-E-semialdehyde decarboxylase (ACMSD) plays an important role in l-tryptophan degradation via the kynurenine pathway. ACMSD forms a homodimer and is functionally inactive as a monomer because its catalytic assembly requires an arginine residue from a neighboring subunit. However, how the oligomeric state and self-association of ACMSD are controlled in solution remains unexplored. Here, we demonstrate that ACMSD from Pseudomonas fluorescens can self-assemble into homodimer, tetramer, and higher-order structures. Using size-exclusion chromatography coupled with small-angle X-ray scattering (SEC-SAXS) analysis, we investigated the ACMSD tetramer structure, and fitting the SAXS data with X-ray crystal structures of the monomeric component, we could generate a pseudo-atomic structure of the tetramer. This analysis revealed a tetramer model of ACMSD as a head-on dimer of dimers. We observed that the tetramer is catalytically more active than the dimer and is in equilibrium with the monomer and dimer. Substituting a critical residue of the dimer-dimer interface, His-110, altered the tetramer dissociation profile by increasing the higher-order oligomer portion in solution without changing the X-ray crystal structure. ACMSD self-association was affected by pH, ionic strength, and other electrostatic interactions. Alignment of ACMSD sequences revealed that His-110 is highly conserved in a few bacteria that utilize nitrobenzoic acid as a sole source of carbon and energy, suggesting a dedicated functional role of ACMSD's self-assembly into the tetrameric and higher-order structures. These results indicate that the dynamic oligomerization status potentially regulates ACMSD activity and that SEC-SAXS coupled with X-ray crystallography is a powerful tool for studying protein self-association.