Electron Transfer Of Redox Proteins Research Articles

Surface-confined redox-active monolayers of a multifunctional anthraquinone derivative on nanoporous and single-crystal gold electrodes

We have investigated self-assembled molecular monolayers (SAMs) of a novel thiol-bearing anthraquinone derivative AQ(OMe) (1-amino-4-((3-mercapto-5-(methoxycarbonyl)phenyl)amino)-9,10-dioxo-9,10-dihydroanthracene-2-sulfonic acid) with multifarious protonation sites, on nanoporous gold (NPG) electrode surfaces. We employ cyclic voltammetry, electrochemical impedance spectroscopy and X-ray photoelectron spectroscopy (XPS). Single-crystal Au(111)-electrode surfaces area used as a reference. XPS exhibits S2p3/2 peaks at 161.1 and 162.1 eV, verifying well-defined AQ(OMe) Au-S bound SAMs. The mid-point potential (E1/2) of the SAMs shifts negatively with increasing pH with a slope of −55 mV pH−1, indicative of a 2e2H redox reaction. The anthraquinone group exhibits quasi-reversible behavior over a wide range of pH values (from2.8to9.0) and a sigmoidal electron transfer (ET) rate constant (kapp) with a maximum value at pH 5.5, which is likely to be close to the isoelectric point. The electrochemically addressable surface coverage, determined from the voltammetric peak areas, decreases with increasing pH. The apparent surface coverage on NPG is more than 13 times higher than that on Au(111), attributed mainly to the larger surface area and different structural packing modes, and possibly also to favorable terrace edges on NPG, but – somewhat unexpectedly – with little effect on kapp. The new anthraquinone derivative offers perspectives for gentle immobilization and efficient mediated interfacial electrochemical ET of redox proteins and other complex biomolecules.

Advances of bioelectrocatalysis by oxidoreductases

<p indent=0mm>Bioelectrocatalysis attracts much attention with the advantages of high selectivity, high efficiency, no byproduct and environmental friendliness. Electrification of industrial biotechnology is challenging for the different operating conditions and catalysts of electrochemical and biochemical reactions. In this review, we present the state-of-the-art information on enzymatic electrosynthesis for the production of fine chemicals. Multidisciplinary strategies of electrode surface modification and enzyme immobilization have been integrated in electrocatalytic devices with various applications in biosensors and bioelectrocatalysis. However, the efficiency of bioelectrosynthesis is determined by the intramolecular electron transfer rate, and interfacial electron transfer rate between the oxidoreductases and electrode. This review aims to summarize recent progresses on the mechanism studies of intramolecular electron transferof redox proteins, including tunneling, hopping, proton-coupled electron transfer, etc. Modular origin of electron transfer chain was also discussed from the perspective of the evolution of spatial adjacency network. Novel and efficient oxidoreductases with enhanced intramolecular and interfacial electron transfer rate can be obtained by protein engineering. The strategies for facilitating intramolecular electron transfer are addressed, which includes the regulation of co-evolved electron transfer network, the designs of molecular switch and the assembly of conductive modules. Oxidoreductases are engineered to improve their biocatalytic performance by using the tools of molecular evolution, modeling, structure prediction, and mutation. Co-evolved molecular switches controll proton-coupled electron transfer and regulate electron transfer inside the multi-center redox proteins. Assembly of surface-binding conductive peptides to oxidoreductases facilitates electricity-driven catalysis. Moreover, the modifications of oxidoreductases allow their predictable immobilization on functionalized electrode surface with improved stability, controlled orientation, surface coverage and enhanced electron transfer. After that, electron transfer within a series of well-defined peptides with orientation-controlled and surface-confined enzymes was addressed. Strategies also developed to increase the biocompatible active surface of electrodes and to promote charge transfer reactions. Nano- and macroporous electrodes based on metal nanoparticles, nanocarbon tubes, graphene, and metallic inverse opals have been designed and fabricated with predictable surface functionalities and electrochemical properties. Electrode/mediator/cofactor/enzyme conjugates can enhance the <italic>in vitro</italic> bioelectricity generation of cofactor. Screening of redox mediators for electrochemical regeneration of NADH was also summarized. Multifunctional surfaces of nano or meso-porous electrodes, where oxidoreductases were bounded to structured electrodes with necessary components (e.g., mediators, cofactor, etc.) with tailored properties, were summarized. Optimizations of functional electrode materials and surfaces can improve the efficiency of cofactor regeneration in the electrochemical reactor modules. Key challenges and future research for better bioelectrocatalysis are discussed and expected briefly. Mining and engineering of novel and robust enzymes from genomic and metagenomic libraries would benefit the bioelectrocatalysis. Based on genome mining, discovery of novel oxidoreductases from extreme microorganism provide thermophilic, alkalophilic, halophilic and organic solvent tolerant oxidoreductases. Reactors for bioelectrocatalysis are optimized to provide a platform for the production of chiral chemicals, which brings great promises for biomanufacturing. Further study of bio-inspired multi-enzyme immobilization which mimics quick electron transfer in multi-enzyme complex is suggested. A variety of approaches for bioelectrocatalysis have been successfully applied for the synthesis of chiral chemicals which are intermediates of fine chemicals and pharmaceuticals. Bioelectrosynthesis processes could replace many chemically catalyzed routes in the future, and aims to construct a new platform for more efficient and green biomanufacturing.

Direct Electrochemical Enzyme Electron Transfer on Electrodes Modified by Self-Assembled Molecular Monolayers

Self-assembled molecular monolayers (SAMs) have long been recognized as crucial “bridges” between redox enzymes and solid electrode surfaces, on which the enzymes undergo direct electron transfer (DET)—for example, in enzymatic biofuel cells (EBFCs) and biosensors. SAMs possess a wide range of terminal groups that enable productive enzyme adsorption and fine-tuning in favorable orientations on the electrode. The tunneling distance and SAM chain length, and the contacting terminal SAM groups, are the most significant controlling factors in DET-type bioelectrocatalysis. In particular, SAM-modified nanostructured electrode materials have recently been extensively explored to improve the catalytic activity and stability of redox proteins immobilized on electrochemical surfaces. In this report, we present an overview of recent investigations of electrochemical enzyme DET processes on SAMs with a focus on single-crystal and nanoporous gold electrodes. Specifically, we consider the preparation and characterization methods of SAMs, as well as SAM applications in promoting interfacial electrochemical electron transfer of redox proteins and enzymes. The strategic selection of SAMs to accord with the properties of the core redox protein/enzymes is also highlighted.

Rationally designed naphthyl substituted amine functionalized ionic liquid platform for covalent immobilization and direct electrochemistry of hemoglobin

Herein, we have designed and demonstrated a facile and effective platform for the covalent anchoring of a tetrameric hemoprotein, hemoglobin (Hb). The platform comprises of naphthyl substituted amine functionalized gel type hydrophobic ionic liquid (NpNH2-IL) through which the heme protein was covalently attached over a glassy carbon electrode (Hb-NpNH2-IL/GCE). UV-vis and FT-IR spectral results confirmed that the Hb on NpNH2-IL retains its native structure, even after being covalently immobilized on NpNH2-IL platform. The direct electron transfer of redox protein could be realized at Hb-NpNH2-IL/GCE modified electrode and a well resolved redox peak with a formal potential of −0.30 V and peak separation of 65 mV was observed. This is due to the covalent attachment of highly conducting NpNH2-IL to the Hb, which facilitates rapid shuttling of electrons between the redox site of protein and the electrode. Further, the fabricated biosensor favoured the electrochemical reduction of bromate in neutral pH with linearity ranging from 12 to 228 µM and 0.228 to 4.42 mM with a detection limit and sensitivities of 3 µM, 430.7 µA mM−1 cm−2 and 148.4 µA mM−1 cm−2 respectively. Notably, the fabricated biosensor showed good operational stability under static and dynamic conditions with high selectivity and reproducibility.

Enhanced direct electron transfer of redox protein based on multiporous SnO2 nanofiber-carbon nanotube nanocomposite and its application in biosensing

A novel third generation H2O2 biosensor is fabricated using multiporous SnO2 nanofiber/carbon nanotubes (CNTs) composite as a matrix for the immobilization of redox protein onto glassy carbon electrode. The multiporous nanofiber (MPNFs) of SnO2 is synthesized by electrospinning technique from the tin precursor. This nanofiber shows high surface area and good electrical conductivity. The SnO2 nanofiber/CNT composite increases the efficiency of biomolecule loading due to its high surface area. The morphology of the nanofiber has been evaluated by scanning electron microscopy (SEM). Cyclic Voltammetry and amperometry technique are employed to study and optimize the performance of the fabricated electrode. A direct electron transfer between the protein's redox centre and the glassy carbon electrode is established after fabrication of the electrode. The fabricated electrode shows excellent electrocatalytic reduction to H2O2. The catalysis currents increases linearly to the H2O2 concentration in a wide range of 1.0 10−6–1.4×10−4M and the lowest detection limit was 30nM (S/N=3). Moreover, the biosensor showed a rapid response to H2O2, a good stability and reproducibility.

Real-Time Dynamic Adsorption Processes of Cytochrome c on an Electrode Observed through Electrochemical High-Speed Atomic Force Microscopy

An understanding of dynamic processes of proteins on the electrode surface could enhance the efficiency of bioelectronics development and therefore it is crucial to gain information regarding both physical adsorption of proteins onto the electrode and its electrochemical property in real-time. We combined high-speed atomic force microscopy (HS-AFM) with electrochemical device for simultaneous observation of the surface topography and electron transfer of redox proteins on an electrode. Direct electron transfer of cytochrome c (cyt c) adsorbed on a self-assembled monolayers (SAMs) formed electrode is very attractive subject in bioelectrochemistry. This paper reports a real-time visualization of cyt c adsorption processes on an 11-mercaptoundecanoic acid-modified Au electrode together with simultaneous electrochemical measurements. Adsorbing cyt c molecules were observed on a subsecond time resolution simultaneously with increasing redox currents from cyt c using EC-HS-AFM. The root mean square roughness (R RMS) from the AFM images and the number of the electrochemically active cyt c molecules adsorbed onto the electrode (Γ) simultaneously increased in positive cooperativity. Cyt c molecules were fully adsorbed on the electrode in the AFM images when the peak currents were steady. This use of electrochemical HS-AFM significantly facilitates understanding of dynamic behavior of biomolecules on the electrode interface and contributes to the further development of bioelectronics.

Direct Electrochemistry and Electrocatalysis of Hemoglobin Immobilized on the Functionalized Graphene-Carbon Nanotube Composite Film

The direct electrochemistry of hemoglobin (Hb) immobilized on the designed carboxyl functionalized polystyrene/graphene-carbon nanotube-Nafion modified indium tin oxide (PS/GE-CNT-Nafion/ITO) electrode and its electrocatalysis for hydrogen peroxide (H2O2) were investigated. UV-visible and Fourier-transform infrared spectra of the resulting Hb-PS/GE-CNT-Nafion composite film suggested that the immobilized Hb retained its original structure. The direct electron transfer behaviors of Hb at the Hb-PS/GE-CNT-Nafion/ITO electrode were investigated by cyclic voltammetry in which a pair of well-defined, quasi-reversible redox peaks with the formal potential (E0′) of −0.249 V (vs. SCE) in phosphate buffer solution (0.05 M, pH 7.0) was obtained. The proposed ITO modified electrode exhibited an excellent electrocatalytic activity to the reduction of H2O2, and the peak currents were linearly related to H2O2 concentration in a wider linearity range from 5.0 × 10−7 to 1.4 × 10−3 M with a correlation coefficient of 0.9983 and a detection limit of 2.0 × 10−7 M (S/N = 3). The sensitivity and the apparent Michaelis-Menten constant () were determined to be 59.1 μA·mM−1 and 8.3 × 10−5 M, respectively. It is expected that the synergetic effects of GE-CNT hybrid materials and functionalized PS provide a general platform for studying of the electron transfer of redox proteins and the design of novel biosensors.

Construction of hyaluronan-silver nanoparticle–hemoglobin multilayer composite film and investigations on its electrocatalytic properties

In this work, hyaluronan-silver nanoparticles (HSNPs) were prepared by UV-initiated photoreduction, and protein hemoglobin (Hb) was then alternately assembled with the prepared negatively charged HSNPs into layer-by-layer (LBL) films on solid surface. The electrochemical behavior and electrocatalytic activities toward oxygen and hydrogen peroxide of the resulting films were studied. It was found that the HSNPs greatly enhanced the electron transfer reactivity of Hb as a bridge. The assembly films showed a pair of nearly reversible redox peaks with a formal potential of −0.32 V (vs. Ag/AgCl) for the heme Fe(III)/Fe(II) redox couple. The immobilized Hb in the films maintained its biological activity, showing a surface-controlled process with a heterogeneous electron transfer rate constant (ks) of 1.0 s−1 and displaying the same features of a peroxidase in the electrocatalytic reduction of oxygen and hydrogen peroxide. This work provides a novel model to fabricate LBL films with protein, polysaccharide and nanoparticles, which may establish a foundation for fabricating new type of biosensors based on the direct electron transfer of redox proteins immobilized in nanocomposite multilayer films with underlying electrodes.

Bicontinuous gyroidal mesoporous carbon matrix for facilitating protein electrochemical and bioelectrocatalytic performances

A strategy of protein-entrapment in bicontinuous gyroidal mesoporous carbon (BGMC) nanocomposite films is described. Herein, the quasi-reversible electron transfer of redox proteins (such as glucose oxidase and myoglobin) is probed and the associated biocatalytic activity is revealed. The apparent heterogeneous electron transfer rate constant of the immobilized glucose oxidase is up to 9.4s−1, much larger than those in carbon nanotubes and some conventional mesoporous carbons. The BGMC based glucose biosensor enables the determination of glucose at a potential of 0.6V (vs. SCE). Its detection limit is 1.0×10−5M (signal-to-noise ratio, S/N=3), the linear response is up to 7.49mM and the detection sensitivity is 52.5nAmM−1 Furthermore, a series of BGMCs with different pore sizes is designed and synthesized using sucrose or phenol formaldehyde resin to study the influences of pore sizes and carbon sources on the immobilization of redox proteins and on the heterogeneous electron transfer.

Layer-by-layer assembly of collagen and electroactive myoglobin

In this work, an electrochemically inert protein collagen was successfully assembled with electroactive myoglobin (Mb) into {collagen/Mb} n layer-by-layer (LbL) films on solid surfaces. UV–vis spectroscopy and cyclic voltammetry (CV) were used to confirm the film growth and characterize the films. The collagen films provided a biocompatible microenvironment for Mb, and Mb in the stable {collagen/Mb} n films showed a nearly reversible CV response for its heme Fe(III)/Fe(II) redox couple. The direct electrochemistry of Mb in the films was used to electrocatalyze the reduction of oxygen and hydrogen peroxide. The interaction between collagen and Mb under different pH conditions was also explored. Not only the positively charged Mb at pH 5.0 but also the negatively charged Mb at pH 9.0 could be assembled with collagen into {collagen/Mb} n films. This work provides a novel example to assemble LbL films with two different proteins, and may establish a foundation for fabricating the new type of biosensors based on the direct electron transfer of redox proteins with underlying electrodes.

Carbon Nanotube‐Based Electrochemical Sensors: Principles and Applications in Biomedical Systems

Carbon nanotubes (CNTs) have received considerable attention in the field of electrochemical sensing, due to their unique structural, electronic and chemical properties, for instance, unique tubular nanostructure, large specific surface, excellent conductivity, modifiable sidewall, high conductivity, good biocompatibility, and so on. Here, we tried to give a comprehensive review on some important aspects of the applications of CNT‐based electrochemical sensors in biomedical systems, including the electrochemical nature of CNTs, the methods for dispersing CNTs in solution, the approaches to the immobilization of functional CNT sensing films on electrodes, and the extensive biomedical applications of the CNT‐based electrochemical sensors. In the last section, we mainly focused on the applications of CNT‐based electrochemical sensors in the analysis of various biological substances and drugs, the methods for constructing enzyme‐based electrochemical biosensors and the direct electron transfer of redox proteins on CNTs. Because several crucial factors (e.g., the surface properties of carbon nanotubes, the methods for constructing carbon nanotube electrodes and the manners for electrochemical sensing applications) predominated the analytical performances of carbon nanotube electrodes, a systematical comprehension of the related knowledge was essential to the acquaintance, mastery and development of carbon nanotube‐based electrochemical sensors.

Electrocatalytic studies of Cytochrome c functionalized single walled carbon nanotubes on self-assembled monolayer of 4-ATP on gold

Direct electron transfer of redox proteins on electrode surfaces using surface immobilized protein films is gaining importance as it provides significant insight into the functioning of biological systems and in the development of biosensors. We have studied functionalized cytochrome c (cyt c) surfaces by covalently using glutaraldehyde as a cross linker and also non-covalently by a simple immersion process of physical adsorption. The third method described in this paper involves combining the affinity of the redox proteins for single walled carbon nanotube (SWNT) surface and its non-covalent binding to the 4-aminothiophenol (4-ATP) self-assembled monolayer on Au. The immobilized composite films facilitate the interfacial electron transfer and electrocatalytic activity of cyt c. The cyt c immobilized surfaces were characterized by scanning probe techniques, cyclic voltammetry, electrochemical impedance spectroscopy and chronoamperometry. The AFM images of the protein immobilized surface show dense distribution of SWNT- cyt c on the surface forming several clusters. A closer examination using STM shows dense distribution of proteins on 4-ATP surface. We have correlated the voltammetric results with the information obtained from AFM and STM images to explain the electrocatalytic activity of the protein layer on the electrode surface.

One-step co-electropolymerized conducting polymer–protein composite film for direct electrochemistry-based biosensors

A novel meliorative conducting polymer–redox protein composite film was fabricated via one-step co-electropolymerization approach. With electrostatic interactions between the conducting polymer and the protein, the composite film possesses attractive features, especially in studies of direct electron transfer of redox proteins as well as the development of unique protein based biosensors. Electrochemical impedance spectroscopy (EIS), atomic force microscopy (AFM), reflectance absorption infrared spectroscopy (RAIR) and ultraviolet visible spectroscopy (UV–vis) were performed to characterize the unique film. The direct electron transfer of the redox protein in the composite film were explored, and its bioactivity was also investigated by catalyzing the reductions of H 2O 2 and NO 2 −, demonstrating its great potential applications in direct electrochemistry-based high performance biosensors.

PH-Dependent Behaviors of Electroactive Myoglobin Loaded into Layer-by-Layer Films Assembled with Alginate and Hydroxyethyl Cellulose Ethoxylate

In the present work, strong polybase quaternized hydroxyethyl cellulose ethoxylate (HECE) and weak polyacid alginate (AA) were assembled into {HECE/AA} n layer-by-layer (LBL) films on electrodes by electrostatic interaction between them, and the films were then immersed in myoglobin (Mb) solution to load Mb into the films, designated as {HECE/AA}n-Mb. The {HECE/AA}n-Mb films showed a nearly reversible cyclic voltammetric (CV) peak pair at about -0.34 V vs SCE in pH 7.0 buffers for Mb heme Fe(III)/Fe(II) redox couple, and the surface concentration of electroactive Mb in the films (Gamma*) was affected significantly by the pH of Mb loading solution and testing solution. The amount of Mb loaded from pH 5.0 solution was much larger than that from pH 9.0 solution, which is mainly attributed to the higher degree of swelling, porosity, and permeability of {HECE/AA}n films at pH 5.0 than at pH 9.0. In addition, the electrostatic interaction between Mb and the AA component in the films might also play an important role in Mb loading. The pH of the testing solution where {HECE/AA}n-Mb films were tested by CV also influenced the Gamma* value, showing that the fraction of electroactive Mb among the total Mb loaded into the films increased remarkably as the pH of the testing solution decreased. This result is rationalized in terms of the pH-dependent film permeability toward counterions and the electron-hopping mechanism in electron transfer of redox proteins in the film phase. This model system may provide a general and effective approach to control the electroactivity of immobilized redox proteins in the multilayer assembly containing weak polyions by adjusting pH and may guide us to develop the new kind of controllable electrochemical biosensors based on the direct electrochemistry of enzymes.

Biosensors based on direct electron transfer in redox proteins

In biosensors based on direct electron transfer in redox proteins, efficient electron-transfer pathways between the immobilized redox protein and the electrode surface have to be established so to allow a fast electron transfer and concomitantly avoiding free-diffusing redox species. In this review, prerequisites for the direct electron transfer of redox proteins and immobilization of redox proteins on the electrode surfaces are addressed. Based on the specific nature of different proteins and non-manual immobilization procedures, possible biosensor designs are discussed, namely biosensors based on (1) ferritin; (2) cytochrome c; (3) myoglobin; (4) hemoglobin; (5) horseradish peroxidase; (6) catalase; (7) glucose oxidase; and (8) xanthine oxidase.

Direct electron transfer of redox proteins on a Nafion-cysteine modified gold electrode

A novel Nafion-cysteine functional membrane was constructed. Rapid and direct electron transfer of horseradish peroxidase, Euphorbia latex amine oxidase, superoxide dismutase and cytochrome c was carried out on the functional membrane modified gold electrode with good stability and repeatability. The immobilized protein modified electrodes showed quasi-reversible electrochemical redox behaviors with formal potentials of 60, 35, 66 and 38 mV (vs. Ag/AgCl) in 20 mM potassium phosphate buffer solution, pH 7.0 at 25 °C. The cathodic transfer coefficients were 0.42, 0.42, 0.45, 0.44 and electron transfer rate constants were evaluated to be 1.2, 1.6, 1.0 and 0.6 s −1.

Direct Electron Transfer of Redox Proteins and Enzymes Promoted by Carbon Nanotube

Direct Electron Transfer of Redox Proteins and Enzymes Promoted by Carbon Nanotube

Proton-coupled electron transfer of cytochrome c.

Cytochrome c (Cyt-c) was electrostatically bound to self-assembled monolayers (SAM) on an Ag electrode, which are formed by omega-carboxyl alkanethiols of different chain lengths (C(x)). The dynamics of the electron-transfer (ET) reaction of the adsorbed heme protein, initiated by a rapid potential jump to the redox potential, was monitored by time-resolved surface enhanced resonance Raman (SERR) spectroscopy. Under conditions of the present experiments, only the reduced and oxidized forms of the native protein state contribute to the SERR spectra. Thus, the data obtained from the spectra were described by a one-step relaxation process yielding the rate constants of the ET between the adsorbed Cyt-c and the electrode for a driving force of zero electronvolts. For C(16)- and C(11)-SAMs, the respective rate constants of 0.073 and 43 s(-1) correspond to an exponential distance dependence of the ET (beta = 1.28 A(-1)), very similar to that observed for long-range intramolecular ET of redox proteins. Upon further decreasing the chain length, the rate constant only slightly increases to 134 s(-1) at C(6)- and remains essentially unchanged at C(3)- and C(2)-SAMs. The onset of the nonexponential distance dependence is paralleled by a kinetic H/D effect that increases from 1.2 at C(6)- to 4.0 at C(2)-coatings, indicating a coupling of the redox reaction with proton-transfer (PT) steps. These PT processes are attributed to the rearrangement of the hydrogen-bonding network of the protein associated with the transition between the oxidized and reduced state of Cyt-c. Since this unusual kinetic behavior has not been observed for electron-transferring proteins in solution, it is concluded that at the Ag/SAM interface the energy barrier for the PT processes of the adsorbed Cyt-c is raised by the electric field. This effect increases upon reducing the distance to the electrode, until nuclear tunneling becomes the rate-limiting step of the redox process. The electric field dependence of the proton-coupled ET may represent a possible mechanism for controlling biological redox reactions via changes of the transmembrane potential.

Correlation of the Structural Decomposition and Performance of Pyridinethiolate Surface Modifiers at Gold Electrodes for the Facilitation of Cytochrome c Heterogeneous Electron-Transfer Reactions

This paper describes the results of an electrochemical and spectroscopic (infrared reflection and X-ray photoelectron spectroscopies) investigation of the modified gold electrode surfaces prepared from dilute ethanolic solutions of 4-mercaptopyridine (PySH) and 4,4‘-dipyridyl disulfide (PySS). Both precursors have been used extensively as facilitators for the electron transfer of redox proteins like cytochrome c (cyt c). During the course of an investigation of the interfacial architectures formed from the two different precursors, a previously unreported structural instability in the adlayers was discovered. This instability manifests itself as a decrease in the ability of the modified surfaces to facilitate the electron transfer of cyt c that correlates with an increase of the immersion time in the precursor solutions. Results are presented that delineate the decrease in facilitator performance and probe the structural changes resulting in the decrease in performance. Together, the electrochemical and surface spectroscopic findings reveal that the modified surfaces spontaneously decompose to yield an adlayer composed largely of adsorbed atomic and oligomeric sulfur, an adlayer that we found to be ineffective in the facilitation of the electron transfer reaction of cyt c. The implications of these findings on the use of this type of modifier to studies of electron transfer reactions of redox proteins and to issues of the general structural stability of organosulfur-based monolayers are briefly discussed.

The direct electrochemistry of cryo-hydrogel immobilized myoglobin at a glassy carbon electrode

A cryo-hydrogel membrane (CHM) immobilized at a glassy carbon (GC) electrode is reported for the direct electron transfer of redox proteins. The most attractive characteristics of this CHM were its hydrophilic micro-environment for incorporated proteins to retain their activities, its high ability for protection against interference of denatured and adsorbed proteins at the electrode, its potential applications for various proteins or enzymes, as well as its high mechanical strength and thermal stability. A clear well developed and stable redox wave was obtained for commercially available horse heart myoglobin without further purification, giving a peak to peak separation ΔE p = 93 mV at 5 mV s −1 and the formal electrode potential E 0′ = −0.158 V (vs. Ag|Agcl). The formal heterogeneous electron transfer rate constant was calculated as k 0′ = 5.7 × 10 −4 cm s −1 at pH 6.5, showing rapid electron transfer was achieved. The pH controlled conformational equilibria, acid state η natural state η basic I state η basic II state, of myoglobin at the CHM GC electrode in the pH range 0–13.8 were also observed and are discussed in detail.