Structural Characterization and Ligand/Inhibitor Identification Provide Functional Insights into the Mycobacterium tuberculosis Cytochrome P450 CYP126A1.
Structural Characterization and Ligand/Inhibitor Identification Provide Functional Insights into the Mycobacterium tuberculosis Cytochrome P450 CYP126A1.
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结构表征和配体/抑制剂鉴定提供了对结核分枝杆菌细胞色素P450 CYP126A1的功能见解。
DOI：
10.1074/jbc.m116.748822
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发表时间：
2017-01-27
期刊：
影响因子：
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通讯作者：
Munro AW
中科院分区：
文献类型：
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作者：
Chenge JT;Duyet LV;Swami S;McLean KJ;Kavanagh ME;Coyne AG;Rigby SE;Cheesman MR;Girvan HM;Levy CW;Rupp B;von Kries JP;Abell C;Leys D;Munro AW
结核分枝杆菌H37Rv基因组编码20种细胞色素P450，包括对感染和细菌活力至关重要的P450。许多M。结核杆菌P450的特征仍然不明，这表明他们的进一步分析可能会提供新的见解M。结核代谢过程和药物发现的新靶点。CYP126A1是广泛分布于分枝杆菌和其他细菌中的P450家族的代表。在这里，我们探讨了CYP126A1的生化和结构特性，包括其与新的化学配体的相互作用。一项关于唑类抗真菌药物的调查显示，CYP126A1被含有咪唑环的唑类药物强烈抑制，但不被含有三唑环的受试药物抑制。为了进一步探索CYP126A1的分子偏好性，寻找酶功能的探针，我们进行了高通量筛选。含有三个或更多个环结构的化合物占主导地位的筛选命中，包括硝基芳香族化合物，诱导底物样的血红素谱的CYP126A1的位移。光谱电化学测量显示，血红素铁电位增加155 mV时，绑定到一个新确定的硝基芳香族药物。在无配体的CYP126A1和与筛选中发现的化合物结合的CYP126A1的晶体结构中观察到CYP126A1二聚体。然而，酮康唑的结合方向破坏了P450二聚体界面处的BC环区域，并导致CYP126A1单体晶体形式。结构数据还显示，硝基芳族配体通过取代CYP126A1远端的水作为底物“月光”，但抑制酶活性。CYP126A1的相对极性活性位点将其与M中最密切相关的甾醇结合P450区分开来。结核病，这表明进一步的研究将揭示其多样的底物选择性。
The Mycobacterium tuberculosis H37Rv genome encodes 20 cytochromes P450, including P450s crucial to infection and bacterial viability. Many M. tuberculosis P450s remain uncharacterized, suggesting that their further analysis may provide new insights into M. tuberculosis metabolic processes and new targets for drug discovery. CYP126A1 is representative of a P450 family widely distributed in mycobacteria and other bacteria. Here we explore the biochemical and structural properties of CYP126A1, including its interactions with new chemical ligands. A survey of azole antifungal drugs showed that CYP126A1 is inhibited strongly by azoles containing an imidazole ring but not by those tested containing a triazole ring. To further explore the molecular preferences of CYP126A1 and search for probes of enzyme function, we conducted a high throughput screen. Compounds containing three or more ring structures dominated the screening hits, including nitroaromatic compounds that induce substrate-like shifts in the heme spectrum of CYP126A1. Spectroelectrochemical measurements revealed a 155-mV increase in heme iron potential when bound to one of the newly identified nitroaromatic drugs. CYP126A1 dimers were observed in crystal structures of ligand-free CYP126A1 and for CYP126A1 bound to compounds discovered in the screen. However, ketoconazole binds in an orientation that disrupts the BC-loop regions at the P450 dimer interface and results in a CYP126A1 monomeric crystal form. Structural data also reveal that nitroaromatic ligands “moonlight” as substrates by displacing the CYP126A1 distal water but inhibit enzyme activity. The relatively polar active site of CYP126A1 distinguishes it from its most closely related sterol-binding P450s in M. tuberculosis, suggesting that further investigations will reveal its diverse substrate selectivity.