Direct electron transfer between copper-containing proteins and electrodes (original) (raw)
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Surface Characterization and Direct Electrochemistry of Multi-Copper Oxidases
A next generation of micro-and nano-scale electronic materials, nano-devices, fuel cells, sensors, and MEMS actuation devices will be based on biocomposite hybrids that combine redox enzymes and conductive materials [1, 2]. Controlling the interfacial organization and understanding intramolecular electron transfer mechanism between the bio-nanostructures and engineered surfaces are crucial for advancing the predicted technologies. The present research aims to advance the understanding of direct electronic communication by using multicopper oxidases and gold or carbon electrodes as experimental platforms. Understanding the biocatalytic energy conversion process will help promote their use in micro-power sources. Multicopper oxidases are an important class of enzymes found across the phylogenetic spectrum. Examples have been identified and characterized from plants, fungi, bacteria and humans. The structure and biochemistry of the multicopper oxidases are reasonably well understood fr...
Electrochimica Acta, 2022
Copper efflux oxidases (CueOs) are key enzymes in copper homeostasis systems. The mechanisms involved are however largely unknown. CueO-type enzymes share a typical structural feature composed of Methionine-rich (Met-rich) domains that are proposed to be involved in copper homeostasis. Bioelectrocatalysis using CueO-type enzymes in the presence of Cu 2+ recently highlighted a new Cu 2+-dependent catalytic pathway related to a cuprous oxidase activity. In this work, we further investigated the effects of Cu 2+ on direct electron transfer (DET)-type bioelectrocatalytic reduction of O2 by CueO at NH2-functionalized multi-walled carbon nanotubes. The DET-type bioelectrocatalytic activity of CueO decreased at low potential in the presence of Cu 2+ , showing unique peak-shaped voltammograms that we attribute to inactivation and reactivation processes. Chronoamperometry was used to kinetically analyze these processes, and the results suggested linear free energy relationships between the inactivation/reactivation rate constant and the electrode potential. Pseudo-steady-state analysis also indicated that Cu 2+ uncompetitively inhibited the enzymatic activity. A detailed model for the Cu 2+-dependent reductive inactivation of CueO was proposed to explain the electrochemical data, and the related thermodynamic and kinetic parameters. A CueO variant with truncated copper-binding α helices and bilirubin oxidase free of Metrich domains also showed such reductive inactivation process, which suggests that multicopper oxidases contain copper-binding sites that lead to inactivation.
Electrochemistry, 2020
We investigated properties of direct electron transfer (DET)-type bioelectrocatalysis of recombinant native Copper efflux oxidase (rCueO) and its variants which lack α helices covering the electron-donating substrate-binding site (Δα5-7CueO, Δα5CueO, Δα6-7CueO, and Δα5-7+1/2α5CueO) at mesoporous carbon electrodes without pretreatment and modified with positively or negatively charged aromatic amines. Kinetic and thermodynamic parameters of the electrode reaction were obtained by analysis of steady-state catalytic waves based on a random orientation model and examined the results on the basis of the structural information of the enzymes. The data suggested that the electron transfer pathway is different from that in solution; electrons are transferred from an electrode to the T1 Cu site through the negatively charged position near the T1 Cu site in rCueO without passing through the α helix region in DET-type bioelectrocatalysis. Positively charged electrode was a suitable scaffold for DET-type reaction of rCueO. The T1 Cu site in Δα5CueO became somewhat hydrophobic and hydrophobic electrode worked as a suitable scaffold for the variant. Negatively charged electrode seems to induce unfavorable attractive orientation for DET-type reaction between the electrode and positively charged region of the CueOs on the opposite side of the T1 Cu site.
Journal of the American Chemical Society, 2014
Cytochrome aa 3 from Paracoccus denitrificans and cytochrome ba 3 from Thermus thermophilus, two distinct members of the heme−copper oxidase superfamily, were immobilized on electrodes modified with gold nanoparticles. This procedure allowed us to achieve direct electron transfer between the enzyme and the gold nanoparticles and to obtain evidence for different electrocatalytic properties of the two enzymes. The pH dependence and thermostability reveal that the enzymes are highly adapted to their native environments. These results suggest that evolution resulted in different solutions to the common problem of electron transfer to oxygen.
Electrochemical and spectral characterization of blue copper protein models
Electrochemistry Communications, 2009
Blue copper proteins play a central role in various enzymatic anabolic/catabolic pathways in living cells by virtue of the integrated metal ions. These ions may exist in variable oxidation states, with suitable reduction potentials and fast electron-transfer rates which in turn is a manifestation of their unusual geometry and coordination. We report the electrochemical and spectral characterization of three novel complexes of copper (II) with N 2 S type tridentate chelating agent 2,2 0-dithiodianiline (dta), having structural similarities to the active site of Type I copper proteins. High positive redox potentials in the range of 0.5-0.6 V vs Ag/AgCl electrode of the complexes and the absorption maxima at $550 nm, with high extinction coefficients, correspond well with typical blue copper proteins. The IR and EPR studies support the assigned pseudo tetrahedral structures to the complexes. The diffusion coefficient and rate constant for heterogeneous charge transfer for Cu 2+ /Cu + coordinated in a potentially bio-mimetic Type I site is reported.
Physical Chemistry Chemical Physics, 2014
The effect of proper enzyme orientation at the electrode surface was explored for two multi-copper oxygen reducing enzymes: Bilirubin Oxidase (BOx) and Laccase (Lac). Simultaneous utilization of ''tethering'' agent (1-pyrenebutanoic acid, succinimidyl ester; PBSE), for stable enzyme immobilization, and syringaldazine (Syr), for enzyme orientation, of both Lac and BOx led to a notable enhancement of the electrode performance. For Lac cathodes tested in solution it was established that PBSE-Lac and PBSE-Syr-Lac modified cathodes demonstrated approximately 6 and 9 times increase in current density, respectively, compared to physically adsorbed and randomly oriented Lac cathodes. Further testing in solution utilizing BOx showed an even higher increase in achievable current densities, thus BOx was chosen for additional testing in air-breathing mode. In subsequent air-breathing experiments the incorporation of PBSE and Syr with BOx resulted in current densities of 0.65 AE 0.1 mA cm À2 ; 2.5 times higher when compared to an unmodified BOx cathode. A fully tethered/oriented BOx cathode was combined with a NAD-dependent Glucose Dehydrogenase anode for the fabrication of a complete enzymatic membraneless fuel cell. A maximum power of 1.03 AE 0.06 mW cm À2 was recorded for the complete fuel cell. The observed significant enhancement in the performance of ''oriented'' cathodes was a result of proper enzyme orientation, leading to facilitated enzyme/electrode interface interactions.
Frontiers in Chemistry, 2018
In recent years, enzymatic fuel cells have experienced a great development promoted by the availability of novel biological techniques that allow the access to a large number of enzymatic catalysts. One of the most important aspects in this area is the development of biocatalysts for the oxygen reduction reaction (ORR). Laccases from the group of enzymes called blue multi-cooper oxidases have received considerable attention because of their ability to catalyze the electrochemical oxygen reduction reaction to water when immobilized on metallic or carbonaceous electrode materials. In this paper we report a comprehensive study of the electrocatalytic activity of the enzyme Copper efflux oxidase (CueO) from Escherichia coli immobilized on different electrode materials. The influence of the electrode substrate employed for protein immobilization was evaluated using glassy carbon, gold or platinum electrodes. Gold and platinum electrodes were modified using different self-assembled monolayers (SAM) able to tune the electrostatic interaction between the protein and the substrate, depending on the nature of the terminal functional group in the SAM. The effects of protein immobilization time, electrode potential, solution pH and temperature, protein and O 2 concentration have been carefully investigated. Finally, direct electron transfer (DET) was investigated in the presence of the following inhibitors: fluoride (F −), chloride (Cl −) and azide (N − 3).
ACS Omega, 2021
This perspective analyzes recent advances in the spectroelectrochemical investigation of redox proteins and enzymes immobilized on biocompatible or biomimetic electrode surfaces. Specifically, the article highlights new insights obtained by surface-enhanced resonance Raman (SERR), surfaceenhanced infrared absorption (SEIRA), protein film infrared electrochemistry (PFIRE), polarization modulation infrared reflection−absorption spectroscopy (PMIRRAS), Forster resonance energy transfer (FRET), X-ray absorption spectroscopy (XAS), electron paramagnetic resonance (EPR), and differential electrochemical mass spectrometry (DMES)-based spectroelectrochemical methods on the structure, orientation, dynamics, and reaction mechanisms for a variety of immobilized species. This includes small heme and copper electron shuttling proteins, large respiratory complexes, hydrogenases, multicopper oxidases, alcohol dehydrogenases, endonucleases, NO-reductases, and dye decolorizing peroxidases, among other enzymes. Finally, I discuss the challenges and foreseeable future developments toward a better understanding of the functioning of these complex macromolecules and their exploitation in technological devices.