Child Stella A, Ghith Amna, Bruning John B, Bell Stephen G
Department of Chemistry, University of Adelaide, SA 5005, Australia.
School of Biological Sciences, University of Adelaide, SA 5005, Australia.
J Inorg Biochem. 2020 Aug;209:111116. doi: 10.1016/j.jinorgbio.2020.111116. Epub 2020 May 20.
The steroid lipid binding cytochrome P450 (CYP) enzymes of Mycobacterium tuberculosis are essential for organism survival through metabolism of cholesterol and its derivatives. The counterparts to these enzymes from Mycobacterium marinum were studied to determine the degree of functional conservation between them. Spectroscopic analyses of substrate and inhibitor binding for the four M. marinum enzymes CYP125A6, CYP125A7, CYP142A3 and CYP124A1 were performed and compared to the equivalent enzymes of M. tuberculosis. The sequence of CYP125A7 from M. marinum was more similar to CYP125A1 from M. tuberculosis than CYP125A6 but both showed differences in the resting heme spin state and in the binding modes and affinities of certain azole inhibitors. CYP125A7 did not show a significant Type II inhibitor-like shift with any of the azoles tested. CYP142A3 bound a similar range of steroids and inhibitors to CYP142A1. However, there were some differences in the extent of the Type I shifts to the high-spin form with steroids and a higher affinity for the azole inhibitors compared to CYP142A1. The two CYP124 enzymes had similar substrate binding properties. M. marinum CYP124 was characterised by X-ray crystallography and displayed strong conservation of active site residues, except near the region where the carboxylate terminus of the phytanic acid substrate would be bound. As these enzymes in M. tuberculosis have been identified as candidates for inhibition the data here demonstrates that alternative strategies for inhibitor design may be required to target CYP family members from distinct pathogenic Mycobacterium species or other bacteria.
结核分枝杆菌的类固醇脂质结合细胞色素P450（CYP）酶通过胆固醇及其衍生物的代谢对生物体存活至关重要。对海分枝杆菌中这些酶的对应物进行了研究，以确定它们之间的功能保守程度。对海分枝杆菌的四种酶CYP125A6、CYP125A7、CYP142A3和CYP124A1进行了底物和抑制剂结合的光谱分析，并与结核分枝杆菌的等效酶进行了比较。海分枝杆菌的CYP125A7序列与结核分枝杆菌的CYP125A1比CYP125A6更相似，但两者在静息血红素自旋状态以及某些唑类抑制剂的结合模式和亲和力方面均存在差异。CYP125A7对所测试的任何唑类均未显示出明显的II型抑制剂样位移。CYP142A3与CYP142A1结合的类固醇和抑制剂范围相似。然而，与CYP142A1相比，类固醇向高自旋形式的I型位移程度存在一些差异，并且对唑类抑制剂的亲和力更高。两种CYP124酶具有相似的底物结合特性。通过X射线晶体学对海分枝杆菌CYP124进行了表征，除了植烷酸底物的羧基末端结合区域附近外，活性位点残基表现出很强的保守性。由于结核分枝杆菌中的这些酶已被确定为抑制的候选对象，此处的数据表明，可能需要采用替代策略来设计抑制剂，以靶向来自不同致病性分枝杆菌物种或其他细菌的CYP家族成员。
