Abstract
We exploit Au/SiO2plasmonic structures to check the effective binding activity of GM1(3) gangliosides hosted in physiological-like biomembranes, in presence of the Epidermal Growth Factor Receptor (EGFR). To this aim, we used bilayers that support the propagation of optical surface plasmon modes (plasmonic transducers, PTs) or guided modes (Plasmon Waveguide Resonators, PWRs). First, we measured the binding of EGFR to GM1(3) by using PTs. Indeed, effective interactions were evidenced, but with faint signals that prevented resolving dissociation kinetics. In order to enhance the optical responses, we turned our attention to PWRs. We first refined the design of a previously adopted Au/SiO2PWR, finding that the nominal sensitivity is independent on SiO2thickness but strongly dependent on its residual losses, due typically to a nonoptimal deposition process. We fabricated an improved Au/SiO2resonator and tested the predicted signal enhancement by monitoring the binding of EGFR to GM3-enriched biomembranes. The measured signal was ~12-fold higher than that one measured using a PT, close to the maximum theoretical enhancement. The higher PWR response enabled us to detect the dissociation of EGFR from GM3, and the value of the apparent dissociation constant of the GM3-EGFR complex could be obtained.
Highlights
Systems mimicking the physiological cell membranes on solid supports are excellent templates for studying in vitro their interactions with several biological molecules
We performed surface plasmon resonance (SPR) analysis with plasmonic transducers (PTs) to evidence the Epidermal Growth Factor (EGF) receptor effective binding with both GM1 and GM3 that resulted more pronounced in the case of Epidermal Growth Factor Receptor (EGFR)-GM3 interaction, in agreement with previous findings
We presented a refined design of the Plasmon Waveguide Resonators (PWRs) device, whose most significant feature is that the maximum theoretical sensitivity (27% nm−1) is quite independent on the SiO2 thickness, suggesting that the control on its growth during the fabrication can be considered as a minor concern
Summary
Systems mimicking the physiological cell membranes on solid supports (solid supported bilayer lipid membranes, ssBLMs) are excellent templates for studying in vitro their interactions with several biological molecules (ionic pumps, signalling agents, etc. . . .). Many of these molecules are recruited in particular mobile areas of the cell membrane, primarily constituted by sphingomyelin (SM) and cholesterol (C) called lipid rafts [1], where microenvironmental effects on their functional properties can occur, with important pharmacological implications [2]. Among these effectors, gangliosides GM1 and GM3 play important roles in many cell functions [3,4,5,6,7]. We completed the characterization evidencing the dissociation of EGFR from the GM3ssBLMs and estimating the apparent dissociation constant KD of the GM3-EGFR complex
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