Distance Dependence of Electron Transfer Kinetics for Azurin Protein Adsorbed to Monolayer Protected Nanoparticle Film Assemblies

Abstract

The distance dependence and kinetics of the heterogeneous electron transfer (ET) reaction for the redox protein azurin adsorbed to an electrode modified with a gold nanoparticle film are investigated using cyclic voltammetry. The nanoparticle films are comprised of nonaqueous nanoparticles, known as monolayer-protected clusters (MPCs), which are covalently networked with dithiol linkers. The MPC film assembly serves as an alternative adsorption platform to the traditional alkanethiolate self-assembled monolayer (SAM) modified electrodes that are commonly employed to study the ET kinetics of immobilized redox proteins, a strategy known as protein monolayer electrochemistry. Voltammetric analysis of the ET kinetics for azurin adsorbed to SAMs of increasing chain length results in quasi-reversible voltammetry with significant peak splitting. We observed rate constants (k degrees (ET)) of 12-20 s(-1) for the protein at SAMs of shorter alkanethiolates that decays exponentially (beta = 0.9/CH(2) or 0.8/A) at SAMs of longer alkanethiolates (9-11 methylene units) or an estimated distance of 1.23 nm and is representative of classical electronic tunneling behavior over increasing distance. Azurin adsorbed to the MPC film platforms of increasing thickness results in reversible voltammetry with very little voltammetric peaks splitting and nearly negligible decay of the ET rate over significant distances up to 20 nm. The apparent lack of distance dependence for heterogeneous ET reactions at MPC film assemblies is attributed to a two-step mechanism involving extremely fast electronic hopping through the MPC film architecture. These results suggest that MPC platforms may be used in protein monolayer electrochemistry to create adsorption platforms of higher architecture that can accommodate greater than monolayer protein coverage and increase the Faradaic signal, a finding with significant implications for amperometric biosensor design and development.

Synthesis, Assembly, and Characterization of Monolayer Protected Gold Nanoparticle Films for Protein Monolayer Electrochemistry

“…As described in prior studies, 2 the original plasmid for the Pseudomonas aeruginosa azurin (AZ) protein was graciously given by Dr. Corey Wilson of Rice University and AZ was provided as a purified and lyophilized powder by the University of Richmond professor, Dr. Jonathan Dattelbaum, that was subsequently rehydrated with 4.4 mM potassium phosphate buffer (KPB, pH = 7.0, μ = 10 mM) to create a 5-10 μM solution as verified by ultraviolet visible (UV-Vis) analysis.…”

“…2,12 This is done by first attaching a MPC film assembled on a 3-MPTMS modified glass slide onto a clean, standard microscope slides using Embed 812 epoxy resin to allow for improved handling during the procedure below. Use caution with the applied heat as higher temperatures will decompose the MPCs within the film.…”

“…Studies have shown that gold nanoparticle film assemblies of this nature provide for a more homogeneous protein adsorption environment and promote ET without distance dependence compared to the more traditional systems modified with alkanethiol self-assembled monolayers (SAM). [1][2][3] In this paper, MPCs functionalized with hexanethiolate ligands are synthesized using a modified Brust reaction 4 and characterized with ultraviolet visible (UV-Vis) spectroscopy, transmission electron microscopy (TEM), and proton ( 1 H) nuclear magnetic resonance (NMR). MPC films are assembled on SAM modified gold electrode interfaces by using a "dip cycle" method of alternating MPC layers and dithiol linking molecules.…”

Direct electrochemistry of horseradish peroxidase immobilized on electrografted 4-ethynylphenyl film via click chemistry

“…The direct, unmediated electrochemistry of redox proteins at electrodes is an important tool for the study of biological electron transfer (ET) processes such as respiration and photosynthesis [1], and the electrodes modified with redox proteins can be potentially applied as biosensors, biocatalysts and biofuel cells [2][3][4]. It is well known that the structural properties inherent in proteins make it difficult to achieve direct ET while the redox proteins are directly immobilized on electrode surfaces.…”

Antimony doped tin oxide nanoparticles and their assembly in mesostructured film

“…Transparent conducting oxides, such as tin-doped indium oxide (ITO) or doped tin oxides, are known as thin dense films, being indispensible for OLEDs or non-silicon solar cells [1,2]. However, other types of optoelectronic devices, especially those involving immobilized species or functional layers, require 3D-conducting networks rather than flat electrode interfaces [3][4][5]. Such 3D-electrode architectures with large interface area are suitable as conducting hosts for the accommodation of functional guest species, leading to a more efficient electron transfer and increased efficiency of the devices.…”