Investigating AHL-lipid bilayer interactions using nonlinear optical spectroscopy

Abstract

Acylated homoserine lactones (AHLs) play an important role in bacterial infection and biofilm formation and also allow communication between microorganisms and their eukaryotic host cells (Inter-kingdom signaling). The detailed mechanism of these interactions is yet unknown. To mimic interactions between AHLs and cell membranes, the incorporation of AHLs with different chain lengths in supported lipid bi- layers (SLBs) was studied using vibrational sum-frequency generation (SFG) spectroscopy. Deuterium- labelled AHLs were synthesized to prevent the overlap of spectroscopic bands allowing the detailed analysis of the interaction. Three AHLs of different chain length were investigated and their orienta- tion within the lipid bilayer was determined. Moreover, the development of AHL-related SFG signals over 5 h was studied providing evidence that AHLs can cross the bilayer in a flip-flop process. However, the determined half life of this process is of the order of days indicating that it is not relevant for the signaling in bacterial biofilms. Furthermore, to relate structural changes with terminal groups and chain lengths, differently ter- minated self-assembled monolayers (SAMs) were studied with SFG spectroscopy in air and in situ. Pyridine-terminated SAMs present a different behavior upon electrochemical desorption depending on an odd or even numbered aliphatic chain. Using SFG spectroscopy it was determined that even numbered pyridine-terminated self-assembled monolayers retain their structure, whereas odd numbered monolayers lose their conformation upon desorption. Furthermore, nonlinear third-order effects were observed with this experimental setup and could be readily explained with theoretical predictions.