Quantitative Analysis of Tethered Vesicle Assemblies by Quartz Crystal Microbalance with Dissipation Monitoring:Binding Dynamics and Bound Water Content

摘要

To implement the molecular recognition properties of membrane proteins for applications including biosensors and diagnostic arrays,the construction of a biomimetic platform capable of maintaining protein structure and function is required.In this paper,we describe a tethered phospholipid vesicle assembly that overcomes the major limitations of planar supported lipid bilayers and alternative biomimetic membrane platforms and characterize it using quartz crystal microbalance with dissipation monitoring (QCM-D) and fluorescence microscopy.We provide evidence of a one-step mechanism for bilayer formation and monitor the subsequent adsorption and binding of streptavidin,vesicles,and streptavidin-coated microspheres.For all three species,we identify a critical surface density above which a significant amount of coupled interstitial water contributes to the response of the quartz resonator in a phenomenon similar to dynamic coupling due to surface roughness.A Sauerbrey-type analysis is sufficient to accurately interpret the QCM-D results for streptavidin binding if water is treated as an additional inertial mass,but viscoelastic models must be invoked for vesicle and microsphere binding.Additionally,we present evidence of vesicle flattening,possibly enhanced by a biotin-mediated membrane-membrane interaction.