Alpha-Helical Peptide-Induced Vesicle Rupture Revealing New Insight into the Vesicle Fusion Process As Monitored in Situ by Quartz Crystal Microbalance-Dissipation and Reflectometry

Abstract

We have used simultaneous quartz crystal microbalance-dissipation (QCM-D) monitoring and four-detector optical reflectometry to monitor in situ the structural transformation of intact vesicles to a lipid bilayer on a gold surface. The structural transformation of lipid vesicles to a bilayer was achieved by introducing a particular amphipathic, alpha-helical (AH) peptide. The combined experimental apparatus allows us to simultaneously follow the acoustic and optical property changes of the vesicle rupturing process upon interaction with AH peptides. While QCM-D and reflectometry have similar sensitivities in terms of mass and thickness resolution, there are unique advantages in operating these techniques simultaneously on the same substrate. These advantages permit us to (1) follow the complex interaction between AH peptides and intact vesicles with both acoustic and optical mass measurements, (2) calculate the amount of dynamically coupled water during the interaction between AH peptides and intact vesicles, (3) demonstrate that the unexpectedly large increase of both adsorbed mass and the film's energy dissipation is mainly caused by swelling of the vesicles during the binding interaction with AH peptides, and (4) permit us to understand the structural transformation from intact vesicles to a bilayer via the AH peptide interaction by monitoring viscoelastic properties, acoustic mass, optical mass, and thickness changes of both the binding and destabilization processes. From the deduced "hydration signature" we followed the complex transformation of lipid assemblies. On the basis of this information, a mechanism of this structural transformation is proposed that provides new insight into the process of vesicle fusion on solid substrates.

A Molecularly Complete Planar Bacterial Outer Membrane Platform

“…Thus, the surface coverage of unruptured OMVs ( θ ) can be estimated as a function of the mass of adsorbed OMVs ( M Ad ), the saturation mass of adsorbed OMVs ( M satd ). and the jamming limit of spheres on a 2D plane (~54%)47:…”

Model Membrane Platforms for Biomedicine: Case Study on Antiviral Drug Development

“…To address this challenge, it is therefore important to characterize biointerfacial processes by using several surface-sensitive techniques that measure different physical principles. To investigate the mechanism by which AH peptide modulates the film properties of the intact vesicle layer, simultaneous QCM-D and reflectometry measurements to follow the AH peptide-mediated vesicle-to-bilayer transformation on gold have been reported [43]. While the acoustic wave-based QCM-D technique characterizes the structural properties of an adlayer film, the optical-based reflectometry technique provides a measure of the bound molecular mass that is related to the effective thickness and the effective refractive index of the adlayer [94].…”

“…6f). An especially useful feature of hydration signature is the direct identification of individual kinetic steps to describe the overall mechanism of biological interactions [43,95]. Taken together, the combination of QCM-D and reflectometry techniques enabled a more complete description of the AH peptide-mediated vesicle rupturing process.…”

“…In this outlook, we introduce areas where we envision engineering strategies may further aid antiviral drug development. By assembling an in situ assay to predict virus particle rupture [40][41][42][43], the work reviewed here has established a cost-effective tool to rapidly screen and functionally characterize antiviral drug candidates in minutes rather than days. Based on this preliminary format, the system can be improved by increasing sensitivity to reduce the amount of peptide material required and/or adapting the signal response to a more readily high-throughput measurement technique.…”

“…As a case study example of a viral pathogen target, we have selected the hepatitis C virus (HCV) and discuss how engineering strategies identified a previously unknown functional activity encoded within the HCV nonstructural 5A (NS5A) protein. By employing a model membrane platform to mimic the host cell membrane interface upon which viral genome replication occurs, a striking ability to lyse lipid vesicles was discovered within the NS5A N-terminal amphipathic, a-helix (AH) [40][41][42][43].…”