Unique Residues at the 3-Fold and 4-Fold Axis of Mycobacterial Ferritin Are Involved in Oligomer Switching
Unique Residues at the 3-Fold and 4-Fold Axis of Mycobacterial Ferritin Are Involved in Oligomer Switching
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DOI：
10.1021/bi301189t
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发表时间：
2013-03-12
期刊：
影响因子：
2.9
通讯作者：
Tyagi, Anil K.
中科院分区：
文献类型：
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作者：
Khare, Garima;Nangpal, Prachi;Tyagi, Anil K.
为了鉴定结核分枝杆菌重要的铁储存蛋白之一BfrB自组装和功能的关键残基，我们采用定点诱变的方法构建了多种突变体。通过对突变体的分析，我们发现了“界面热点残基”（R69、L129和F159），它们是BfrB寡聚化的“开关点”，我们的观察显示了4重轴残基在组装形成中的重要性。此外，我们证明单点突变Q51A， Q126A和E135A可以增强蛋白质的热稳定性，而不影响其组装。重要的是，对各种突变的比较分析表明，不同铁蛋白中不同同源位置的功能可能存在差异；因此，仅仅基于序列结构比较来预测残基的功能可能是不合适的。因此，我们在BfrB的组装形成和功能中发现了新的残基，并表明单点突变具有改变铁蛋白多种特性的显著潜力。此外，本研究发现的“开关残基”或“界面热点”也可以为界面抑制剂的合理设计提供帮助。
To identify the crucial residues involved in the self-assembly and function of BfrB, one of the important iron storage proteins of Mycobacterium tuberculosis, we constructed various mutants by employing site-directed mutagenesis. The analysis of mutants led to the identification of "interface hot-spot residues" (R69, L129, and F159) that act as "switch points" for BfrB oligomerization, and our observations show the importance of 4-fold axis residues in assembly formation. Moreover, we demonstrate that single point mutations Q51A, Q126A, and E135A can enhance the thermal stability of the protein without affecting its assembly. Importantly, a comparative analysis of various mutations revealed that the function of various homologous positions in different ferritins could be at variance; hence, predicting the function of a residue just based on sequence structure comparisons may not be appropriate. Thus, we report the identification of novel residues in the assembly formation and function of BfrB and show that single point mutations have a remarkable potential for alteration of multiple properties of ferritins. Besides, "switch residues" or "interface hot spots" identified in this study could also prove to be helpful for the rational design of interfacial inhibitors.