Silica nanoparticles for the layer-by-layer assembly of fully electro-active cytochrome c multilayers.

Sven C Feifel,Fred Lisdat

doi:10.1186/1477-3155-9-59

Abstract

BackgroundFor bioanalytical systems sensitivity and biomolecule activity are critical issues. The immobilization of proteins into multilayer systems by the layer-by-layer deposition has become one of the favorite methods with this respect. Moreover, the combination of nanoparticles with biomolecules on electrodes is a matter of particular interest since several examples with high activities and direct electron transfer have been found. Our study describes the investigation on silica nanoparticles and the redox protein cytochrome c for the construction of electro-active multilayer architectures, and the electron transfer within such systems. The novelty of this work is the construction of such artificial architectures with a non-conducting building block. Furthermore a detailed study of the size influence of silica nanoparticles is performed with regard to formation and electrochemical behavior of these systems.ResultsWe report on interprotein electron transfer (IET) reaction cascades of cytochrome c (cyt c) immobilized by the use of modified silica nanoparticles (SiNPs) to act as an artificial matrix. The layer-by-layer deposition technique has been used for the formation of silica particles/cytochrome c multilayer assemblies on electrodes. The silica particles are characterized by dynamic light scattering (DLS), Fourier transformed infrared spectroscopy (FT-IR), Zeta-potential and transmission electron microscopy (TEM). The modified particles have been studied with respect to act as an artificial network for cytochrome c and to allow efficient interprotein electron transfer reactions. We demonstrate that it is possible to form electro-active assemblies with these non-conducting particles. The electrochemical response is increasing linearly with the number of layers deposited, reaching a cyt c surface concentration of about 80 pmol/cm2 with a 5 layer architecture. The interprotein electron transfer through the layer system and the influence of particle size are discussed.ConclusionsThis study demonstrates the ability to construct fully electro-active cyt c multilayer assemblies by using carboxy-modified silica nanoparticles. Thus it can be shown that functional, artificial systems can be build up following natural examples of protein arrangements. The absence of any conductive properties in the second building block clearly demonstrates that mechanisms for electron transfer through such protein multilayer assemblies is based on interprotein electron exchange, rather than on wiring of the protein to the electrode.The construction strategy of this multilayer system provides a new controllable route to immobilize proteins in multiple layers featuring direct electrochemistry without mediating shuttle molecules and controlling the electro-active amount by the number of deposition steps.

Highlights

Introduction of the carboxyl groups ontoSiO2-NH2 particle surface After the amino groups were grafted onto the SiO2 particle surface, they need to be coupled with succinc acid anhydride to introduce the carboxyl groups onto the surface
The absence of any conductive properties in the second building block clearly demonstrates that mechanisms for electron transfer through such protein multilayer assemblies is based on interprotein electron exchange, rather than on wiring of the protein to the electrode
It is demonstrated that polymer-free cyt c multilayer assemblies can be built up

Summary

Introduction

Introduction of the carboxyl groups ontoSiO2-NH2 particle surface After the amino groups were grafted onto the SiO2 particle surface, they need to be coupled with succinc acid anhydride to introduce the carboxyl groups onto the surface. Our study describes the investigation on silica nanoparticles and the redox protein cytochrome c for the construction of electro-active multilayer architectures, and the electron transfer within such systems. Raising the number of layers a continuous increase of the voltammetric cyt c signal has been achieved with this system and can be used for the detection of superoxide radicals with enhanced sensitivity [30,31] Such multilayer assemblies have recently been shown to enable incorporation of enzymes and establish communication to the electrode, allowing the construction of different analytical signal chains [24,32,33]. SiNPs have already been used in biosensors since the high surface area of nano-sized silica particles can increase the surface molecule loading and lead to a higher performance of the biosensor [25,34,35,36]

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Conclusion