Catalytic Activity and Proton Translocation of Reconstituted Respiratory Complex I Monitored by Surface-Enhanced Infrared Absorption Spectroscopy

• Published in: Langmuir 2018-05, Vol.34 (20), p.5703-5711
• Main Authors: Gutiérrez-Sanz, Oscar, Forbrig, Enrico, Batista, Ana P, Pereira, Manuela M, Salewski, Johannes, Mroginski, Maria A, Götz, Robert, De Lacey, Antonio L, Kozuch, Jacek, Zebger, Ingo
• Format: Article
• Language: English
• Subjects: Catalysis; Electron Transport Complex I - chemistry; Electron Transport Complex I - metabolism; NAD - chemistry; Oxidation-Reduction; Protons; Spectrophotometry, Infrared
• ISSN: 0743-7463; 1520-5827
• DOI: 10.1021/acs.langmuir.7b04057

Respiratory complex I (CpI) is a key player in the way organisms obtain energy, being an energy transducer, which couples nicotinamide adenine dinucleotide (NADH)/quinone oxidoreduction with proton translocation by a mechanism that remains elusive so far. In this work, we monitored the function of CpI in a biomimetic, supported lipid membrane system assembled on a 4-aminothiophenol (4-ATP) self-assembled monolayer by surface-enhanced infrared absorption spectroscopy. 4-ATP serves not only as a linker molecule to a nanostructured gold surface but also as pH sensor, as indicated by concomitant density functional theory calculations. In this way, we were able to monitor NADH/quinone oxidoreduction-induced transmembrane proton translocation via the protonation state of 4-ATP, depending on the net orientation of CpI molecules induced by two complementary approaches. An associated change of the amide I/amide II band intensity ratio indicates conformational modifications upon catalysis which may involve movements of transmembrane helices or other secondary structural elements, as suggested in the literature [ Di Luca , Proc. Natl. Acad. Sci. U.S.A., 2017, 114, E6314−E6321 ].