pH-dependent thermodynamic intermediates of pHLIP membrane insertion determined by solid-state NMR spectroscopy

Abstract

-glycerol-3-phosphocholine unilamellar vesicles as the model membrane, we systematically examined the state of pHLIP-membrane interactions (in terms of the membrane locations of D/E residues, as well as lipid dynamics) at the intervening pH values of 6.4, 6.1, and 5.8, along with the known states at pH 7.4 and 5.3. Thermodynamic intermediate states distinct from the initial and end states were discovered to exist at each of the intervening pH examined. They support a multistage model of pHLIP insertion in which the D/E titrations occur in a defined sequence at distinct intermediate pH values. This multistage model has important ramifications in pHLIP applications.