Delivering Transmembrane Peptide Complexes to the Gas Phase Using Nanodiscs and Electrospray Ionization

The gas-phase conformations of dimers of the channel-forming membrane peptide gramicidin A (GA), produced from isobutanol or aqueous solutions of GA-containing nanodiscs (NDs), are investigated using electrospray ionization-ion mobility separation-mass spectrometry (ESI-IMS-MS) and molecular dynamics (MD) simulations. The IMS arrival times measured for (2GA + 2Na)²⁺ ions from isobutanol reveal three different conformations, with collision cross-sections (Ω) of 683 Å² (conformation 1, C1), 708 Å² (C2), and 737 Å² (C3). The addition of NH₄CH₃CO₂ produced (2GA + 2Na)²⁺ and (2GA + H + Na)²⁺ ions, with Ω similar to those of C1, C2, and C3, as well as (2GA + 2H)²⁺, (2GA + 2NH₄)²⁺, and (2GA + H + NH₄)²⁺ ions, which adopt a single conformation with a Ω similar to that of C2. These results suggest that the nature of the charging agents, imparted by the ESI process, can influence dimer conformation in the gas phase. Notably, the POPC NDs produced exclusively (2GA + 2NH₄)²⁺ dimer ions; the DMPC NDs produced both (2GA + 2H)²⁺ and (2GA + 2NH₄)²⁺ dimer ions. While the Ω of (2GA + 2H)²⁺ is similar to that of C2, the (2GA + 2NH₄)²⁺ ions from NDs adopt a more compact structure, with a Ω of 656 Å². It is proposed that this compact structure corresponds to the ion conducting single stranded head-to-head helical GA dimer. These findings highlight the potential of NDs, combined with ESI, for transferring transmembrane peptide complexes directly from lipid bilayers to the gas phase. Graphical Abstract ᅟ.