Crystal structure and properties of CYP231A2 from the thermoacidophilic Archaeon Picrophilus torridus
Crystal structure and properties of CYP231A2 from the thermoacidophilic Archaeon Picrophilus torridus
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DOI：
10.1021/bi702240k
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发表时间：
2008-02-19
期刊：
影响因子：
2.9
通讯作者：
Poulos, Thomas L.
中科院分区：
文献类型：
--
作者：
Ho, Winny W.;Li, Huiying;Poulos, Thomas L.
来自嗜热嗜酸菌 Picrophilus torridus 的细胞色素 P450 CYP231A2 (PTO1399) 的晶体结构已被解析。该结构揭示了一个宽阔的基底进入通道。为了更好地了解 CYP231A2 中配体诱导的结构转变，使用 4-苯基咪唑研究了蛋白质-配体相互作用。比较无配体和结合配体的 CYP231A2 结构显示构象变化，其中 F 和 G 螺旋作为单个刚体围绕 F 螺旋 N 末端的枢轴点摆动，使 F 螺旋区域向血红素倾斜，从而与配体更紧密地接触。热熔解数据表明，配体结合后，CYP231A2 的熔解温度增加了近 10 摄氏度，从而表明闭合构象更加稳定。此外，光谱数据表明活性位点在 pH 4.5 下是稳定的，尽管铁的硫醇配体不同寻常地可以可逆质子化。 CYP231A2 不表现出通常与嗜热蛋白相关的结构特征，例如盐桥网络的增加或广泛的芳香族簇。相反，与其他 P450 相比，热稳定性的提高与 CYP231A2 中较小的尺寸和较短的环最相关。
The crystal structure of a cytochrome P450 from the thermoacidophile Picrophilus torridus, CYP231A2 (PTO1399), has been solved. This structure reveals a wide open substrate access channel. To better understand ligand-induced structural transitions in CYP231A2, protein-ligand interactions were investigated using 4-phenylimidazole. Comparison of the ligand-free and -bound CYP231A2 structures shows conformational changes where the F and G helices swing as a single rigid body about a pivot point at the N-terminal end of the F helix, allowing the F helix region to dip toward the heme, resulting in closer contacts with the ligand. Thermal melting data illustrate that the melting temperature for CYP231A2 increases nearly 10 degrees C upon ligand binding, thus illustrating that the closed conformation is substantially more stable. Furthermore, spectroscopic data indicate that the active site is stable at pH 4.5, although, unusually, the thiolate ligand to the iron can be reversibly protonated. CYP231A2 does not exhibit structural features normally associated with thermophilic proteins such as an increase in salt bridge networks or extensive aromatic clustering. The increase in thermal stability instead is best correlated with the smaller size and shorter loops in CYP231A2 compared to other P450s.