Roles of semiquinone species in proton pumping mechanism by complex I.
Roles of semiquinone species in proton pumping mechanism by complex I.
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DOI：
10.1007/s10863-014-9557-9
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发表时间：
2014-08
影响因子：
3
通讯作者：
Sakyiama, Joseph A.
中科院分区：
文献类型：
--
作者：
Nakamaru-Ogiso, Eiko;Narayanan, Madhavan;Sakyiama, Joseph A.
关键词：
复合物I（NDH-1）利用电子转移能量使质子跨膜易位。两种不同的耦合机制目前正在讨论复杂的I：直接（氧化还原驱动）和间接（构象驱动）。半醌（SQ）中间体被认为是耦合机制的关键。最近，利用渐进功率饱和和模拟技术，通过对E. coli复合物I重组为蛋白脂质体。在解偶联剂短杆菌肽D或强效短杆菌肽E存在下，快速松弛SQ（SQNf）信号完全消失。大肠杆菌复合物I抑制剂鳞霉素。慢松弛SQ（SQNs）信号对短杆菌肽D不敏感，但对鳞霉素敏感。极慢松弛SQ（SQNvs）信号对短杆菌肽D和鳞霉素均不敏感。有趣的是，在ΔNuoL突变体中没有观察到SQN信号，该突变体缺乏转运蛋白模块亚基NuoL和NuoM。此外，我们寻求了使用甲基萘醌（它具有较低的氧化还原电位相比，泛醌）作为电子受体的质子泵的化学计量的效果，通过在体外重建实验与泛醌丰富或甲基萘醌丰富的双敲除膜囊泡，其中既不包含复合物I也不NDH-2（非质子易位NADH脱氢酶）。在ΔNuoL和D178 N突变体中没有观察到甲萘醌和泛醌之间的质子泵送化学计量的差异，这被认为缺乏间接质子泵送机制。然而，在野生型中，甲基萘醌的质子泵送化学计量减少了一半。讨论了SQ中间体在配合物I偶联机制中的作用和关系。
Complex I (NDH-1) translocates protons across the membrane using electron transfer energy. Two different coupling mechanisms are currently being discussed for complex I: direct (redox-driven) and indirect (conformation-driven). Semiquinone (SQ) intermediates are suggested to be key for the coupling mechanism. Recently, using progressive power saturation and simulation techniques, three distinct SQ species were resolved by EPR analysis of E. coli complex I reconstituted into proteoliposomes. The fast-relaxing SQ (SQNf) signals completely disappeared in the presence of the uncoupler gramicidin D or the potent E. coli complex I inhibitor squamotacin. The slow-relaxing SQ (SQNs) signals were insensitive to gramicidin D, but they were sensitive to squamotacin. The very slow-relaxing SQ (SQNvs) signals were insensitive to both gramicidin D and squamotacin. Interestingly, no SQNs signal was observed in the ΔNuoL mutant, which lacks transporter module subunits NuoL and NuoM. Furthermore, we sought out the effect of using menaquinone (which has a lower redox potential compared to that of ubiquinone) as an electron acceptor on the proton pumping stoichiometry by in vitro reconstitution experiments with ubiquinone-rich or menaquinone-rich double knock-out membrane vesicles, which contain neither complex I nor NDH-2 (non-proton translocating NADH dehydrogenase). No difference in the proton pumping stoichiometry between menaquinone and ubiquinone was observed in the ΔNuoL and D178N mutants, which are considered to lack the indirect proton pumping mechanism. However, the proton pumping stoichiometry with menaquinone decreased by half in the wild-type. The roles and relationships of SQ intermediates in the coupling mechanism of complex I are discussed.
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