Melittin

All peptides were generated commercially using solid-phase peptide synthesis. Melittin was produced by GenScript (Piscataway, NJ), while Pro-P1 and Pro-P2 were produced by American Peptide Company (Sunnyvale, CA).

Expression and purification of HAA has been described earlier [32 (link)]. The HAB clone (generous gift from Prof. J. Horwitz, Jules Stein Eye Institute, UCLA), was expressed and purified according to Horwitz et al [33 (link)]. Melittin was obtained from Genscript (Piscataway, NJ), and used without further purification.

1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) and 1,2-dimyristoyl-sn-glycero-3-phospho-(1’-rac-glycerol) (DMPG) lipids were purchased from Avanti Polar Lipids. LL-37 was purchased from Bachem. Gramicidin A was purchased from Alfa Aesar. Melittin was purchased from GenScript. Ammonium acetate, Amberlite XAD-2, and sodium cholate were purchased from Sigma Aldrich. Membrane scaffold protein MSP1D1(−) was expressed in E. coli and purified by immobilized metal affinity chromatography (IMAC) as previously described.^{16 (link),17 (link)}Nanodiscs were assembled without peptides as previously described.^{16 (link),17 (link)} Briefly, DMPC or DMPG lipids in chloroform were dried under nitrogen and solubilized in sodium cholate. MSP1D1(−) was added to the lipids, and detergent was removed by addition of Amberlite XAD-2 hydrophobic beads. Nanodiscs were purified using a Superose 6 Increase 10/300 column (GE Healthcare) with 0.2 M Ammonium acetate at pH 6.8.^{18 (link)}