Deletion of a unique loop in the mycobacterial F-ATP synthase subunit sheds light on its inhibitory role in ATP hydrolysis-driven H+ pumping
Deletion of a unique loop in the mycobacterial F-ATP synthase subunit sheds light on its inhibitory role in ATP hydrolysis-driven H+ pumping
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DOI：
10.1111/febs.13715
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发表时间：
2016-05-01
期刊：
影响因子：
5.4
通讯作者：
Grueber, Gerhard
中科院分区：
文献类型：
--
作者：
Hotra, Adam;Suter, Manuel;Grueber, Gerhard
F1FO-ATP合成酶是满足分枝杆菌生命周期中增殖需氧和缺氧休眠阶段能量需求所必需的酶之一。大多数F-ATP合成酶在(3):(3)头部消耗ATP来驱动亚基，亚基通过亚基底部将ATP裂解与F-O c环的质子泵送耦合。atp酶驱动的H+泵送在分枝杆菌中是潜伏的。Priya R等人（2013）J bioenergy Biomembr 45,121 -129在这里，我们使用快速生长的耻辱分枝杆菌的倒置膜囊泡（IMVs）和各种共价和非共价抑制剂来表征IMVs内F-ATP合成酶的ATP水解活性。这些囊泡形成了一个平台，通过删除耻垢分枝杆菌基因组中各自的环编码序列（(166-179)）来研究独特的分枝杆菌环的功能。在含有（166-179）突变蛋白的imv中观察到ATP水解驱动的H+泵送，而在含有野生型F-ATP合酶的imv中没有观察到。此外，与野生型酶相比，含有（166-179）突变蛋白的imv显示出ATP切割增加和ATP合成水平降低，表明该环影响ATP酶活性、ATP酶驱动的H+泵送和ATP合成。这些结果进一步表明，该环可能以不同的模式影响ATP水解和合成的偶联。
The F1FO-ATP synthase is one of the enzymes that is essential to meet the energy requirement of both the proliferating aerobic and hypoxic dormant stages of the life cycle of mycobacteria. Most F-ATP synthases consume ATP in the (3):(3) headpiece to drive the subunit, which couples ATP cleavage with proton pumping in the c ring of F-O via the bottom of the subunit. ATPase-driven H+ pumping is latent in mycobacteria. The presence of a unique 14 amino acid residue loop of the mycobacterial subunit has been described and aligned in close vicinity to the c-ring loop Priya R et al. (2013) J Bioenerg Biomembr 45, 121-129 Here, we used inverted membrane vesicles (IMVs) of fast-growing Mycobacterium smegmatis and a variety of covalent and non-covalent inhibitors to characterize the ATP hydrolysis activity of the F-ATP synthase inside IMVs. These vesicles formed a platform to investigate the function of the unique mycobaterial loop by deleting the respective loop-encoding sequence ((166-179)) in the genome of M. smegmatis. ATP hydrolysis-driven H+ pumping was observed in IMVs containing the (166-179) mutant protein but not for IMVs containing the wild-type F-ATP synthase. In addition, when compared to the wild-type enzyme, IMVs containing the (166-179) mutant protein showed increased ATP cleavage and lower levels of ATP synthesis, demonstrating that the loop affects ATPase activity, ATPase-driven H+ pumping and ATP synthesis. These results further indicate that the loop may affect coupling of ATP hydrolysis and synthesis in a different mode.