Re-examination of inhibitor resistance conferred by Qo-site mutations in cytochrome b using yeast as a model system
Re-examination of inhibitor resistance conferred by Qo-site mutations in cytochrome b using yeast as a model system
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DOI：
10.1002/ps.1066
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发表时间：
2005-10-01
影响因子：
4.1
通讯作者：
Meunier, B
中科院分区：
文献类型：
--
作者：
Fisher, N;Meunier, B
来自酵母（Saccharomyces cerevisiae Meyer ex汉森酵母）的细胞色素B为研究来自多种生物体的Q（0）位点抑制剂（QoI）抗性突变提供了方便的模型系统。来自真菌植物病原体的QoI抗性突变将疟原虫（G143 A和F129 L）、疟原虫（Y279 C/S）和卡氏肺孢子虫（L275 F）（一种人类条件致病真菌）导入酵母细胞色素B中，研究了它们对多种天然细胞色素结合的影响。监测BC（1）复合物的合成（阿托伐醌、嘧菌酯和吡唑醚菌酯）抑制剂。还重新检查了L275 S（来自粘噻唑耐药酵母）。Stigmatellin结合相对不受这些突变的引入。观察到嗜球果伞素类抑制剂粘噻唑、嘧菌酯和吡唑醚菌酯的抗性显著增加，其中G143 A赋予的抗性增加最大。相反，阿托伐醌结合受Y279 C/S和L275 S影响最大。值得注意的是，F129 L、G143 A和L275 S对bc（1）活性的影响很小，因此不太可能在体内产生显著的适应性惩罚。这些数据是根据最近得到的粘噻唑和嘧菌酯抑制的牛bc（1）的原子结构进行讨论的。我们提出，由于G143 A的QoI电阻产生的抑制剂和细胞色素B之间的空间位阻，而其他突变的电阻的机制是由于蛋白质和抑制剂分子之间的结合能增加。定点诱变也用于模拟酵母细胞色素B中哺乳动物Q（0）位点的选定区域，以进一步了解这些QoI在哺乳动物和病原体bc（1）复合物中的不同功效。(c)2005化学工业协会。
Cytochrome b from yeast (Saccharomyces cerevisiae Meyer ex Hansen) provides a convenient model system for the study of Q(0)-site inhibitor (QoI) resistance mutations from a variety of organisms. QoI resistance mutations from fungal plant pathogens (G143A and F129L), malaria agent Plasmodium sp (Y279C/S), and Pneumocystis carinii (L275F), an opportunistic pathogenic fungus of man, were introduced into yeast cytochrome b and their effect on the binding of a variety of natural (myxothiazol and stigmatellin) and synthetic (atovaquone, azoxystrobin and pyraclostrobin) inhibitors to the bc(1) complex monitored. L275S (from a myxothiazol-resistant yeast) was also re-examined. Stigmatellin binding was relatively unaffected by the introduction of these mutations. Significant increases in resistance were observed for the strobilurin-class inhibitors myxothiazol, azoxystrobin and pyraclostrobin, with the largest increase in resistance conferred by G143A. In contrast, atovaquone binding was most effected by Y279C/S and L275S. Notably, F129L, G143A and L275S had a minor effect on bc(1) activity, and so are unlikely to confer significant fitness penalties in vivo. These data are discussed in the light of the atomic structures for myxothiazol- and azoxystrobin-inhibited bovine bc(1) which have recently become available. We propose that QoI resistance due to G143A arises from steric hindrance between the inhibitor and cytochrome b, whereas the mechanism of resistance for the other mutations is due to an increase in binding energy between the protein and inhibitor molecule. Site-directed mutagenesis was also used to model selected regions of the mammalian Q(0) site in yeast cytochrome b in order to further understand the differential efficacy of these QoI in the mammalian and pathogen bc(1) complexes. (c) 2005 Society of Chemical Industry.