Siebels Ilka, Dröse Stefan
Molecular Bioenergetics Group, Medical School, Johann Wolfgang Goethe-University, 60590, Frankfurt am Main, Germany; Goethe University Frankfurt, Institute of Organic Chemistry and Chemical Biology, Buchmann Institute for Molecular Life Sciences, Protein Reaction Control Group, Max-von-Laue-Str. 15, 60438, Frankfurt am Main, Germany.
Molecular Bioenergetics Group, Medical School, Johann Wolfgang Goethe-University, 60590, Frankfurt am Main, Germany; Department of Anesthesiology, Intensive-Care Medicine and Pain Therapy, University Hospital Frankfurt, 60590, Frankfurt am Main, Germany.
Biochem Biophys Res Commun. 2016 Oct 14;479(2):277-282. doi: 10.1016/j.bbrc.2016.09.059. Epub 2016 Sep 14.
The charge translocation by purified reconstituted mitochondrial complex I from the obligate aerobic yeast Yarrowia lipolytica was investigated after adsorption of proteoliposomes to solid-supported membranes. In presence of n-decylubiquinone (DBQ), pulses of NADH provided by rapid solution exchange induced charge transfer reflecting steady-state pumping activity of the reconstituted enzyme. The signal amplitude increased with time, indicating 'deactive→active' transition of the Yarrowia complex I. Furthermore, an increase of the membrane-conductivity after addition of 5-(N-ethyl-N-isopropyl)amiloride (EIPA) was detected which questiones the use of EIPA as an inhibitor of the Na/H-antiporter-like subunits of complex I. This investigation shows that electrical measurements on solid-supported membranes are a suitable method to analyze transport events and 'active/deactive' transition of mitochondrial complex I.
将来自专性需氧酵母解脂耶氏酵母的纯化重组线粒体复合物I吸附到固体支持膜上后，对其电荷转运进行了研究。在存在正癸基泛醌（DBQ）的情况下，通过快速溶液交换提供的NADH脉冲诱导电荷转移，反映了重组酶的稳态泵送活性。信号幅度随时间增加，表明解脂耶氏复合物I发生了“失活→激活”转变。此外，检测到添加5-（N-乙基-N-异丙基）阿米洛利（EIPA）后膜电导率增加，这对将EIPA用作复合物I的Na/H反向转运体样亚基的抑制剂提出了质疑。这项研究表明，在固体支持膜上进行电学测量是分析线粒体复合物I的转运事件和“激活/失活”转变的合适方法。
