V-ATPase is an ATP-driven rotary motor that vectorially transports ions. Together with F-ATPase, a homologous protein, several models on the ion transport have been proposed, but their molecular mechanisms are yet unknown. V-ATPase from Enterococcus hirae forms a large supramolecular protein complex (total molecular weight: -700,000) and physiologically transports Na+ and Li+ across a hydrophobic lipid bilayer. Stabilization of these cations in the binding site has been discussed on the basis of X-ray crystal structures of a membrane-embedded domain, the K-ring (Na+ and Li+ bound forms). Sodium or lithium ion binding-induced difference FTIR spectra of the intact E. hirae V-ATPase have been measured in aqueous solution at physiological temperature. The results suggest that sodium or lithium ion binding induces the deprotonation of Glu139, a hydrogen-bonding change in the tyrosine residue and rigid a-helical structures. Identical difference FTIR spectra between the entire V-ATPase complex and K-ring strongly suggest that protein interaction with the I subunit does not cause large structural changes in the K-ring. This result supports the previously proposed Na+ transport mechanism by V-ATPase stating that a flip-flop movement of a carboxylate group of Glu139 without large conformational changes in the K-ring accelerates the replacement of a Na+ ion in the binding site. This article is part of a Special Issue entitled: Vibrational spectroscopies and bioenergetic systems. (C) 2014 Elsevier B.V. All rights reserved.