Functional Studies on Membrane Proteins by Means of H/D Exchange in Infrared: Structural Changes in Na+ NQR from V. cholerae in the Presence of Lipids
H/D exchange kinetics at the level of the amide proton in the mid infrared (1700–1500 cm−1) make it possible to study the conformational flexibility of membrane proteins, independent of size or the presence of detergent or lipids. Slow, medium, and fast exchanging domains are distinguished, which reveal a different accessibility to the solvent. Whereas amide hydrogens undergo rapid exchange with solvent in an open structure, hydrogens experience much slower exchange when involved in H-bonded structures or when sterically inaccessible to the solvent. Here, we describe the protocol that was used to study the effect of phospholipids on the overall structure of the Na+ NQR from V. cholerae, a sodium pumping membrane protein.
Key wordsH/D exchange kinetics Secondary structure analysis Infrared spectroscopy
We are grateful to the University of Strasbourg, the CNRS, and the foundation FRC for ongoing financial support.
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