Structure of the ferrous form of (4-hydroxyphenyl)pyruvate dioxygenase from Streptomyces avermitilis in complex with the therapeutic herbicide, NTBC
Structure of the ferrous form of (4-hydroxyphenyl)pyruvate dioxygenase from Streptomyces avermitilis in complex with the therapeutic herbicide, NTBC
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DOI：
10.1021/bi049317s
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发表时间：
2004-06-01
期刊：
影响因子：
2.9
通讯作者：
Moran, GR
中科院分区：
文献类型：
--
作者：
Brownlee, JM;Johnson-Winters, K;Moran, GR
（4-羟基苯基）丙酮酸双加氧酶（HPPD）二酮和三酮抑制剂是一种有效的除草剂和治疗药物。抑制活性用于停止植物和人类中亲脂性氧化还原辅助因子的产生，以防止因酪氨酸分解代谢的特定先天缺陷而产生的有毒代谢副产物的积累。在2.5埃的分辨率下，测定了阿维菌链霉菌与抑制剂2-[2-硝基-4-（三氟甲基）苯甲酰]1,3-环己二酮（NTBC）配合物中Fe-II型HPPD的三维结构。NTBC与活性位点金属离子相对应，金属离子位于蛋白质c端结构域中一个可溶解的宽腔的底部。铁主要以五坐标、扭曲的方锥体排列，其中Glu349、His187和His270是蛋白质衍生的配体，另外两个配体来自NTBC的5‘和7’氧。在其中一个原聚体的第6配位位点存在一个低占用水分子。到His270的距离在2.5埃处异常长，其取向从理想的配体几何形状略微扭曲到抑制剂硝基的2.8埃范围内。与其他细菌来源的HPPD的四聚四元结构相比，不对称单元由两个弱相关的原聚物组成，埋表面积为1266埃(2)，共有12个氢键，没有静电相互作用。整个三级结构与来自荧光假单胞菌的HPPD相似（Serre et al., (1999) structure 7,977 -988），尽管c端α -螺旋的位置发生了显著变化。这个c端α -螺旋提供了Phe364，它与Phe336结合，夹住了结合的NTBC的苯环；没有观察到其他显著的氢键或电荷对相互作用。此外，结构表明，除了Val 189外，NTBC只与完全保守的氨基酸发生接触。双齿金属离子配位和pi堆叠芳香环的结合暗示了底物的结合模式和/或过渡状态，这可能是这些抑制剂对HPPD具有极高亲和力的原因。
Di- and triketone inhibitors of (4-hydroxyphenyl)pyruvate dioxygenase (HPPD) are both effective herbicides and therapeutics. The inhibitory activity is used to halt the production of lipophilic redox cofactors in plants and also in humans to prevent accumulation of toxic metabolic byproducts that arise from specific inborn defects of tyrosine catabolism. The three-dimensional structure of the Fe-II form of HPPD from Streptomyces avermitilis in complex with the inhibitor 2-[2-nitro-4-(triflouromethyl)benzoyl]1,3-cyclohexanedione (NTBC) has been determined at a resolution of 2.5 Angstrom. NTBC coordinates to the active site metal ion, located at the bottom of a wide solvent-accessible cavity in the C-terminal domain of the protein. The iron is liganded in a predominantly five-coordinate, distorted square-pyramidal arrangement in which Glu349, His187, and His270 are protein-derived ligands and two other ligands are from the 5' and 7' oxygens of NTBC. There is a low-occupancy water molecule in the sixth coordination site in one of the protomers. The distance to His270 is unusually long at 2.5 Angstrom, and its orientation is somewhat distorted from ideal ligand geometry to within 2.8 Angstrom of the inhibitor nitro group. In contrast to the tetrameric quartenary structure observed for HPPD from other bacterial sources, the asymmetric unit is composed of two weakly associated protomers with a buried surface area of 1266 Angstrom(2) and a total of 12 hydrogen-bonding and no electrostatic interactions. The overall tertiary structure is similar to that of HPPD from Pseudomonas fluorescens (Serre et al., (1999) Structure 7, 977-988), although the position of the C-terminal alpha-helix is dramatically shifted. This C-terminal alpha-helix provides Phe364, which in combination with Phe336 sandwiches the phenyl ring of the bound NTBC; no other significant hydrogen-bonding or charge-pairing interactions are observed. Moreover, the structure reveals that, with the exception of Val 189, NTBC makes contacts to only fully conserved amino acids. The combination of bidentate metal-ion coordination and pi-stacked aromatic rings is suggestive of a binding mode for the substrate and/or a transition state, which may be the origin of the exceedingly high affinity these inhibitors have for HPPD.