Visualizing the movement of the amphipathic helix in the respiratory complex I using a nitrile infrared probe and SEIRAS

Abstract

Conformational movements play an important role in enzyme catalysis. Respiratory complex I, an L-shaped enzyme, connects electron transfer from NADH to ubiquinone in its peripheral arm with proton translocation through its membrane arm by a coupling mechanism still under debate. The amphipathic helix across the membrane arm represents a unique structural feature. Here, we demonstrate a new way to study conformational changes by introducing a small and highly flexible nitrile infrared (IR) label to this helix to visualize movement with surface-enhanced IR absorption spectroscopy. We find that labeled residues K551CL and Y590CL move to a more hydrophobic environment upon NADH reduction of the enzyme, likely as a response to the reorganization of the antiporter-like subunits in the membrane arm.