Exchange of Gramicidin between Lipid Bilayers: Implications for the Mechanism of Channel Formation.

The canonical mechanism of gramicidin (gA) channel formation is transmembrane dimerization of nonconducting subunits that reside in opposite bilayer leaflets. The channels do not open and close; they appear and disappear due to subunit association and dissociation. Many different types of experiments support this monomer ↔ dimer mechanism. Recently, however, this mechanism was challenged, based on experiments with lipid vesicle-incorporated gA under conditions where vesicle fusion could be controlled. In these experiments, sustained channel activity was observed long after fusion had been terminated, which led to the proposal that gA single-channel current transitions result from closed-open transitions in long-lived bilayer-spanning dimers. This proposal is at odds with 40 years of experiments, but involves the key assumption that gA monomers do not exchange between bilayers. We tested the possibility of peptide exchange between bilayers using three different types of experiments. First, we demonstrated the exchange of gA between 1,2-dierucoyl-sn-glycero-3-phosphocholine (DC