Interkingdom Signaling: Integration, Conformation, and Orientation of N-Acyl-l-homoserine Lactones in Supported Lipid Bilayers

Abstract

N-Acyl-l-homoserine lactones (AHLs) are small cell-to-cell signaling molecules involved in the regulation of population density and local gene expression in microbial communities. Recent evidence shows that contact of this signaling system, usually referred to as quorum sensing, to living eukaryotes results in interactions of AHL with host cells in a process termed "inter-kingdom signaling". So far details of this process and the binding site of the AHLs remain unknown; both an intracellular and a membrane-bound receptor seem possible, the first of which requires passage through the cell membrane. Here, we used sum-frequency-generation (SFG) spectroscopy to investigate the integration, conformation, orientation, and translocation of deuterated N-Acyl-L-homoserine lactones (AHL-dn) with varying chain length (8, 12, and 14 C atoms) in lipid bilayers consisting of a 1:1 mixture of POPC:POPG supported on SiO2 substrates (prepared by vesicle fusion). We found that all AHL-dn derivatives are well-ordered within the supported lipid bilayer (SLB) in a preferentially all-trans conformation of the deuterated alkyl chain and integrated into the upper leaflet of the SLB with the methyl terminal groups pointing downwards. For the bilayer system described above, no flip-flop of AHL-dn from the upper leaflet to the lower one could be observed. Spectral assignments and interpretations were further supported by Fourier transform infrared and Raman spectroscopy.

Synthesis and analysis of stable isotope-labelled N-acyl homoserine lactones

“…These signals can be used to study the integration of deuterated AHLs in supported bilayers. 29,58 To demonstrate and further extend the possible applications of these deuterated compounds, two AHL-degradation products, which occur in nature, were synthesized. The hydrolysis of the lactone ring of AHLs towards N-acyl homoserines, occurs in a temperature, pH and chain length dependent way and results in a loss of QS-activity.…”

Assessing the Gold Standard: The Complex Vibrational Nonlinear Susceptibility of Metals

“…In the process, we have provided a route for determining ' S for any sample, dielectric or metal. 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 ...…”

“…The nonresonant response from gold provides the background signal that interferes constructively with the ODT methyl response to provide the upwards-pointing peaks, and destructively with the response from the octadecanol methyl group resulting in downwards-pointing peaks. The versatility of this method is evidenced by the range of molecules and interfaces it has since been applied to, providing phase information from polymer-metal interfaces, 18 lipid bilayers and cell structures, [19][20][21] monolayers, [22][23][24][25][26] and immobilized proteins. 17,27 The above method for phase detection allows the relative phase of the resonant modes to be determined.…”

Unique Vibrational Features as a Direct Probe of Specific Antigen–Antibody Recognition at the Surface of a Solid‐Supported Hybrid Lipid Bilayer

“…More specifically, second‐order NLO techniques have set one′s heart on biological interfaces for multiple reasons, first of all the possibility to univocally probe those nano‐bio‐interfaces while surrounded by a macroscopic liquid environment, with the unique advantage that the latter does not contribute to the signal . For example, lipid mono‐ or bilayers mimicking cell membranes are still extensively investigated in interaction with peptides or membrane proteins . In addition to fundamental research purposes, second‐order NLO spectroscopies have provided successful proofs in detecting biological recognition in nanoscale devices.…”

“…[9,10,11] For example, lipid mono-or bilayers mimicking cell membranes are still extensively investigated in interaction with peptides or membrane proteins. [12][13][14][15][16][17][18][19][20][21][22][23][24][25] In addition to fundamental research purposes, second-order NLO spectroscopies have provided successful proofs in detecting biological recognition in nanoscale devices. Second harmonic generation (SHG) ands um frequency generation (SFG) probe the electronic and vibrational responses, respectively,o f interfacial systems.…”