Measuring melittin binding to planar supported lipid bilayer by chiral second harmonic generation Original Research Article

Second harmonic generation (SHG) has been used for the label-free detection of melittin upon intercalation into planar-supported lipid bilayers of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphotidylcholine (POPC). A novel counter-propagating SHG optical geometry was used to effectively isolate the achiral and chiral contributions of the surface, thus allowing for an investigation of both binding and structural contributions to melittin adsorption. Melittin binding to the membrane was monitored as a function of bulk concentration through detection of both the chiral and achiral SHG signal. Analysis of the SHG adsorption isotherms revealed Langmuir adsorption behavior for the achiral SH emission and Frumkin adsorption behavior for the chiral SH emission. The binding constants (Ka) obtained for the chiral and achiral SHG responses were determined to be 8.3±1.0×105 and 1.68±0.40×107 M−1 respectively. The frequency dependent SHG response was also obtained for the achiral and chiral contributions. The SHG spectra show good correlation with the spectral transitions observed in both the UV–VIS and CD spectra of melittin in solution. The results of these studies have far reaching implications in the use of SHG for the analysis of protein interfacial phenomena.