Electron Transfer Reactions Of Cytochrome Research Articles

<i>In situ</i> Observation of Direct Electron Transfer Reaction of Cytochrome <i>c</i> Immobilized on ITO Electrode Modified with 10-carboxydecylphosphonic Acid by Slab Optical Waveguide Spectroscopy and Cyclic Voltammetry

<i>In situ</i> Observation of Direct Electron Transfer Reaction of Cytochrome <i>c</i> Immobilized on ITO Electrode Modified with 10-carboxydecylphosphonic Acid by Slab Optical Waveguide Spectroscopy and Cyclic Voltammetry

Electron-Transfer Reaction of Cytochrome c Adsorbed on Mixed Alkanethiol Monolayer Electrode and Its Electrocatalytic Activity

Electron-Transfer Reaction of Cytochrome c Adsorbed on Mixed Alkanethiol Monolayer Electrode and Its Electrocatalytic Activity

OS7-3-3 スラブ光導波路分光法を用いたITO電極上に吸着したチトクロ-ムcの直接電子移動反応のその場観察(OS7 最先端材料が拓くマイクロ・ナノ工学の新展開(3))

OS7-3-3 スラブ光導波路分光法を用いたITO電極上に吸着したチトクロ-ムcの直接電子移動反応のその場観察(OS7 最先端材料が拓くマイクロ・ナノ工学の新展開(3))

Computer simulation and SERR detection of cytochrome c dynamics at SAM-coated electrodes

In this paper we present a combined experimental and theoretical study of the heterogeneous electron transfer reaction of cytochrome c electrostatically adsorbed on metal electrodes coated with monolayers of 6-mercaptohexanoic acid. Molecular dynamics simulations and pathways calculations show that adsorption of the protein leads to a broad distribution of orientations and, thus, to a correspondingly broad distribution of electron transfer rate constants due to the orientation-dependence of the electronic coupling parameter. The adsorbed protein exhibits significant mobility and, therefore, the measured reaction rate is predicted to be a convolution of protein dynamics and tunnelling probabilities for each orientation. This prediction is confirmed by time-resolved surface enhanced resonance Raman which allows for the direct monitoring of protein (re-)orientation and electron transfer of the immobilised cytochrome c. The results provide a consistent explanation for the non-exponential distance-independence of electron transfer rates usually observed for proteins immobilized on electrodes.

Electrochemistry of Mitochondria: A New Way to Understand Their Structure and Function

AbstractIn this article, electrochemistry of mitochondria is achieved. Cyclic voltammograms of freshly prepared mitochondria were obtained by immobilizing mitochondria together with glutaraldehyde and bovine serum albumin on the surface of a pyrolytic graphite electrode. Two pairs of redox peaks could be observed which were ascribed to the electron transfer reactions of cytochrome c and FAD/FADH2. Study of submitochondrial particles was also conducted, which could confirm the results of the study of the entire mitochondria. The redox wave of NADH could be obtained due to the destruction of the membrane of mitochondria. We have also checked the function of succinate in mitochondria by employing the electrochemical method. This work is not only the first to be able to obtain the direct electrochemistry of mitochondria, but is also beneficial to the further understanding of the structure and function of mitochondria in vitro.

Kinetics of the electron transfer reaction of Cytochrome c 552 adsorbed on biomimetic electrode studied by time-resolved surface-enhanced resonance Raman spectroscopy and electrochemistry

Cytochrome c (552) (Cyt-c (552)) and its redox partner ba ( 3 )-oxidase from Thermus thermophilus possess structural differences compared with Horse heart cytochrome c (cyt-c)/cytochrome c oxidase (CcO) system, where the recognition between partners and the electron transfer (ET) process is initiated via electrostatic interactions. We demonstrated in a previous study by surface-enhanced resonance Raman (SERR) spectroscopy that roughened silver electrodes coated with uncharged mixed self-assembled monolayers HS-(CH(2))( n )-CH(3)/HS-(CH(2))( n + 1)-OH 50/50, n = 5, 10 or 15, was a good model to mimic the Cyt-c (552) redox partner. All the adsorbed molecules are well oriented on such biomimetic electrodes and transfer one electron during the redox process. The present work focuses on the kinetic part of the heterogeneous ET process of Cyt-c (552) adsorbed onto electrodes coated with such mixed SAMs of different alkyl chain length. For that purpose, two complementary methods were combined. Firstly cyclic voltammetry shows that the ET between the adsorbed Cyt-c (552) and the biomimetic electrode is direct and reversible. Furthermore, it allows the estimation of both the density surface coverage of adsorbed Cyt-c (552) and the kinetic constants values. Secondly, time-resolved SERR (TR-SERR) spectroscopy showed that the ET process occurs without conformational change of the Cyt-c (552) heme group and allows the determination of kinetic constants. Results show that the kinetic constant values obtained by TR-SERR spectroscopy could be compared to those obtained from cyclic voltammetry. They are estimated at 200, 150 and 40 s(-1) for the ET of Cyt-c (552) adsorbed onto electrodes coated with mixed SAMs HS-(CH(2))( n )-CH(3)/HS-(CH(2))( n + 1)-OH 50/50, n = 5, 10 or 15, respectively.

Interactions between Cytochrome c and DNA Strands Self-Assembled at Gold Electrode

In this work, we reported the investigation on the interaction between DNAstrands self-assembled at gold electrodes and an electron transfer protein, cytochrome c. Weobserved that cytochrome c exhibited well-defined electrochemistry in both double-strandedand single-stranded DNA films. This suggested that the electron transfer reaction ofcytochrome c arose possibly due to the electron hopping along DNA strands rather thanwiring along the double helix. We also compared the heterogeneous electron transfer rate ofcytochrome c with that of a ruthenium complex, which further confirmed this mechanism.

Electron Transfer Reactions of Cytochrome c Using the Mixed Monolayer Au Electrode Composed of CoIII Complex and Hydroxyl-Terminated Alkanethiols

We prepared the mixed monolayer-modified Au electrode composed of negatively charged CoIII complex (1) and hydroxyl-terminated alkanethiols (Cn; n = 2, 6), 1/Cn-Au, using the ‘step-by-step’ immobilization method; the low-density monolayer of 1 (1-Au) was firstly constructed with a spacing and then Cn molecules were filled in the gap of 1. In the cyclic voltammetric measurement of horse heart cytochrome c (cyt c) using 1/C2-Au, only a pair of oxidation-reduction wave was observed with almost the same peak separation as the case of 1-Au. Since the electron transfer rate by using the single monolayer of C2 was much faster than that of 1-Au, it clearly indicates that the modification of C2 does not cause the phase-separation and C2 molecules on Au do not influence to the electron transfer reaction with cyt c. In contrast, 1/C6-Au showed faster electron transfer rate than the cases of 1-Au and 1/C2-Au, but exhibited slower rate than that of single monolayer of C6. These findings indicate that cyt c firstly interacts with negatively charged 1, and then electron transfer reaction proceeds via C6 molecules on Au electrode.

Importance of poly(ethylene oxide)-modification and chloride anion for the electron transfer reaction of cytochrome c in 1-ethyl-3-methylimidazolium bis(trifluoromethanesulfonyl)imide

Horse heart cytochrome c (cyt c) was chemically modified with poly(ethylene oxide) (PEO) to dissolve it in room temperature ionic liquid 1-ethyl-3-methylimidazolium bis(trifluoromethanesulfonyl)imide ([emim][TFSI]). The redox response of the modified cyt c, hereafter PEO–cyt c, was analyzed in [emim][TFSI]. PEO modification to the surface of cyt c, which exceeded 60% of the total mass of the PEO–cyt c, was an effective method to solubilize the cyt c. In spite of the high ion density and sufficient ionic conductivity of [emim][TFSI], no redox response of pure PEO–cyt c was detected. However, a reversible redox response of PEO–cyt c was observed after adding a simple electrolyte such as KCl to [emim][TFSI]. The redox response of PEO–cyt c was sensitive to the anion radius of the added salt, and the chloride anion was found to be the best anion species to produce a redox response of PEO–cyt c in [emim][TFSI]. However, above a certain salt concentration, the resulting increase in solution viscosity would suppress the redox reaction. The results strongly indicate that the chloride anions, because of their mobility in the polypeptide matrix, compensate the charge change of heme during the electron transfer reaction. Larger anions did not show such an effect due to sterical restrictions on the migration through the protein shell to the heme pocket of cyt c.

Direct electrochemistry of engineered cytochrome b562 molecules with a ligand binding pocket

The rapid and reversible electron transfer reaction of cytochrome b 562 was observed at an In 2O 3 electrode. The estimated heterogeneous electron transfer rate constant ( k 0′) was k 0′ ≥ 5.0 × 10 −3 cm s −1 at pH 6.5. When the methionine-7 (Met-7) residue, which coordinates to the heme iron as an axial ligand, of the wild-type cytochrome b 562 was replaced by an Ala or Gly residue, a water molecule bound to the heme iron and the electron transfer rate constants decreased to 1.3 × 10 −3 and 1.8 × 10 −3 cm s −1, respectively. This decrease in the electron transfer rate would be due to the larger reorganization energy for the structural change at the redox site. The midpoint potential of cytochrome b 562 was shifted negatively by ∼135 mV by replacing Met-7 with Ala or Gly. Similar dissociation kinetics of cyanide for the mutated molecules as compared to native myoglobin was obtained.

Dynamics of Electron Transfer Pathways in Cytochrome c Oxidase

Cytochrome c oxidase mediates the final step of electron transfer reactions in the respiratory chain, catalyzing the transfer between cytochrome c and the molecular oxygen and concomitantly pumping protons across the inner mitochondrial membrane. We investigate the electron transfer reactions in cytochrome c oxidase, particularly the control of the effective electronic coupling by the nuclear thermal motion. The effective coupling is calculated using the Green's function technique with an extended Huckel level electronic Hamiltonian, combined with all-atom molecular dynamics of the protein in a native (membrane and solvent) environment. The effective coupling between Cu A and heme a is found to be dominated by the pathway that starts from His B204 . The coupling between heme a and heme a 3 is dominated by a through-space jump between the two heme rings rather than by covalent pathways. In the both steps, the effective electronic coupling is robust to the thermal nuclear vibrations, thereby providing fast and efficient electron transfer.

The reaction of cytochrome c from different species with cytochrome c oxidase immobilized in an electrode supported lipid bilayer membrane

In past work the direct electron transfer reactions of bovine cytochrome c oxidase in an electrode-supported lipid bilayer membrane have been studied. Its reaction with cytochrome c in solution was also studied and found to be consistent with previous solution studies. In this work it is shown that the electron transfer reactions of cytochrome c oxidase in this electrode-supported lipid bilayer membrane depend on the source of cytochrome c. This property has also been widely studied for solution samples. The differences in the electron transfer reaction rates correlate with the differences in the amino acid sequence for the cytochrome c molecules studied. Electrochemical results suggest that the dissociation of the cytochrome c/cytochrome c oxidase reaction complex is the rate-controlling step in this electron transfer mechanism for cytochrome c from some sources. Moreover, the electron transfer reaction mechanism exhibits biphasic reaction kinetics, which is consistent with earlier work on reactions between solubilized cytochrome c oxidase/cytochrome c samples. These results indicate that the cytochrome c oxidase modified electrodes described herein could be used to distinguish amino acid sequence variations in proteins such as cytochrome c, and this has potential relevance as a diagnostic for disease states.

Electron Transfer Reaction of Cytochrome c at the Electrode Surface Analyzed with Noncontact Optical Waveguide Spectroscopy

Electron Transfer Reaction of Cytochrome c at the Electrode Surface Analyzed with Noncontact Optical Waveguide Spectroscopy

Kinetic studies on the oxidation of cytochrome b(5) Phe35 mutants with cytochrome c, plastocyanin and inorganic complexes.

To illustrate the functions of the aromatic residue Phe35 of cytochrome b(5) and to give further insight into the roles of the Phe35-containing hydrophobic patch and/or aromatic channel of cytochrome b(5), we studied electron transfer reactions of cytochrome b(5) and its Phe35Tyr and Phe35Leu variants with cytochrome c, with the wild-type and Tyr83Phe and Tyr83Leu variants of plastocyanin, and with the inorganic complexes [Fe(EDTA)](-), [Fe(CDTA)](-) and [Ru(NH(3))(6)](3+). The changes at Phe35 of cytochrome b(5) and Tyr83 of plastocyanin do not affect the second-order rate constants for the electron transfer reactions. These results show that the invariant aromatic residues and aromatic patch/channel are not essential for electron transfer in these systems.

The [Ru(CN)5(pyS)]4- Complex, an Efficient Self-Assembled Monolayer for the Cytochrome c Heterogeneous Electron Transfer Studies

The organothiol 4-mercaptopyridine (pyS) has been used extensively as facilitator for the assessment of heterogeneous electron transfer reaction of cytochrome c (cyt c). Its efficiency, however, is strongly affected by the instability of the adlayer due to the C-S bond cleavage. The K(4)[Ru(CN)(5)(pyS)].3H(2)O complex was synthesized and characterized aiming its utilization as an inorganic self-assembled monolayer (SAM) that would enhance the gold adlayer stability. The SAM formed by this complex onto gold (RupySAu) was characterized by spectroscopic (FTIRRAS and SERS) and electrochemical (LSV) techniques. The ex situ vibrational SERS and FTIRRAS spectra data of this SAM formed onto gold suggest a sigma interaction between the gold and sulfur atoms of the complex, inducing a perpendicular arrangement in relation to the surface normal. Additionally, SERS and FTIRRAS spectra performed for freshly prepared RupySAu adlayer and for large immersion times in the precursor solution have not shown any significant change that would reflect the degradation of the adlayer. The LSV desorption curves of this SAM indicate an enhancement in the C-S bond strength of the pyS ligand when coordinated to the [Ru(CN)(5)](3-) moiety. Comparatively to the data obtained for the desorption process of the pyS monolayer, the reductive desorption potential, E(rd), of the RupySAu presents a shift of -17 mV. This bond strength intensification leads to an increase in the stability of the monolayer. The voltammetric curves of cyt c carried out with the RupySAu electrode showed electrochemical parameters consistent with those reported for the native protein, as well as the maintenance of the electrochemical kinetic data after repetitive cycles. The results all together suggest that the pi back-bonding effect from the [Ru(CN)(5)](3-) metal center plays an important role in the stability of the RupySAu adlayer, improving the assessment of the cyt c heterogeneous electron transfer reaction.

Novel Time-Resolved Surface-Enhanced (Resonance) Raman Spectroscopic Technique for Studying the Dynamics of Interfacial Processes: Application to the Electron Transfer Reaction of Cytochrome c at a Silver Electrode

A novel approach for time-resolved (TR) surface-enhanced (resonance) Raman (SE(R)R) spectroscopy is presented for probing potential-dependent processes of molecules adsorbed on a silver electrode. TR SE(R)R spectroscopy offers the unique advantage of providing structural and kinetic data exclusively of the adsorbed species and their reactions. These processes are initiated by a rapid potential jump and monitored by SE(R)R spectroscopy after a delay time δ for the probe interval Δ t. The synchronization is achieved by a mechanical chopper that triggers the potential jump via a photodiode and gates the exciting continuous-wave laser beam. After the probe event, the potential is reset to its initial value. Thus, the original equilibrium is restored to allow a continuous repetition of the sequence of potential jumps and probe events. During the entire experiment, the detection system, a liquid nitrogen-cooled charge-coupled device (CCD) detector, is active so that the signal-to-noise ratio (SNR) can be iteratively improved. This mode of detection does not limit the time resolution, so that the present approach allows TR SE(R)R experiments down to the microsecond time scale without lowering the SNR. The possibilities and limitations of this method are discussed. As an example we present preliminary results of a TR SERR study on yeast iso-1 cytochrome c (Cyt- c) adsorbed on a Ag electrode by applying a potential jump from −0.4 V to +0.05 V (vs. saturated calomel electrode). The experiments are carried out with a rotating electrode to avoid photoinduced degradation and desorption processes. The SERR spectra, which were measured with delay times between 45 to 175 ms, were analyzed quantitively in terms of the various states of the adsorbed Cyt- c that are formed in this potential range. The results show that under these conditions the relaxation processes include the electron transfer of the adsorbed Cyt- c to the electrode and a subsequent conformational transition. The analysis of the data reveals a heterogenous oxidation rate constant of 10.3 s−1 and rate constant for the conformational transition of 4.3 s−1, supporting the view that the biological electron transfer of Cyt- c is coupled with conformational transitions.

Photosensitized electron transfer reactions of cytochrome c4 from Pseudomonas stutzeri with flavins and methyl viologen

Rate constants for electron transfer reactions of the fully reduced, fully oxidized, and half-reduced forms of the di-heme protein cytochrome c 4 from Pseudomonas stutzeri with several flavin species and methyl viologen were determined by laser flash absorption spectroscopy. The flavin reactions include both triplet and photoreduced semiquinone forms. The rate constant values range from 0.7 × 10 7 to 1.6 × 10 9M −1 s −1. The dependence on ionic strength for flavin-mononucleotide semiquinone and methyl viologen follows the behaviour expected from reactions with negatively charged protein sites. Considering these data and those previously reported for electron transfer between cytochrome c 4 and the redox couples [Co(terpy) 2] 2x/3y and [Co(bipy) 3] 2x/3y (terpy=2,2′,2″-terpyridine; bipy=2,2′-bipyridine), a free energy relation spanning of 0.85 cV was calculated. The total reorganization Gibbs free energy inferred from that relation ( E R=1–1.2 eV) exceeds the value for electron transfer between several single-heme proteins and flavin species (∼0.7–0.9 eV). This could reflect the spin changes associated with electron transfer reactions of cytochrome c 4. The rate constants for cytochrome c 4 at the given driving force are two to five times larger than for single-heme proteins. This implies that the electronic transmission coefficient is about two orders of magnitude larger than for the single-heme electron transfer processes.

Electron Transfer Reaction of Cytochrome c at Poly(ethylene oxide)-Thiolate-Modified Gold Electrode

Abstract Gold electrode was modified with poly(ethylene oxide)s having thiol end (PEO-SH). The electron transfer reaction of cytochrome c was evaluated by the gold electrode modified with PEO-SH, having PEO molecular weight of 150, where the adsorption of proteins and resulting denaturation were restrained. The increase of PEO molecular weight conducted suppression of the redox responses at the modified gold electrode.

Photoinduced electron transfer in singly labeled thiouredopyrenetrisulfonate cytochrome c derivatives.

A novel method for initiation of intramolecular electron transfer reactions in cytochrome c is reported. The method is based on photoexcitation of covalently attached thiouredopyrenetrisulfonate (TUPS) by the third harmonic frequency of a Nd:YAG laser (355 nm), the reaction that generates the low-potential triplet state of the dye with high quantum efficiency. TUPS derivatives of horse heart cytochrome c singly labeled at specific lysine residues were prepared and purified to homogeneity by ion-exchange high-pressure liquid chromatography. Eight derivatives were characterized by determination of the location of the modification, reduction potentials, and measurement of enzymatic activity with cytochrome oxidase. Transient absorption spectroscopy was used to directly measure the rate constants for the electron transfer reaction from the photoexcited triplet state of TUPS to the oxidized heme group and the back reaction from the ferrous heme to the oxidized dye. For all singly labeled derivatives, the rate constants for heme reduction were 1 or 2 orders of magnitude larger than for its reoxidation, consistent with the greater thermodynamic driving force for the oxidation reaction. Analysis of the variation of electron-transfer rates with the distance separating the dye and the heme reveals a value of coupling decay constant (beta) of 0.46 A-1. Rapid and effective photoreduction of cytochrome c makes it a useful tool for fast initiation of electron transfer in the reductive direction within complexes of cytochrome c with other redox proteins.

Electron tunneling in proteins: role of the intervening medium

Analysis of electron-transfer (ET) kinetics data obtained from experiments on Ru-modified proteins (cytochrome c, azurin, myoglobin) reveals that distant donor-acceptor electronic couplings depend upon the secondary structure of the intervening polypeptide matrix. Rates of Fe2+→Ru3+ ET reactions in cytochrome c decay exponentially with tunneling-pathway length (decay constant 0.73 A–1); these rates also decay exponentially with Ru-Fe distance (decay constant 1.1 A–1). In azurin, a β-sheet protein, Cu+→Ru3+ rates exhibit an exponential Cu-Ru distance dependence with a decay constant of 1.1 A–1. Comparison of distant couplings in azurin and myoglobin suggests that hydrogen bonds are better mediators across β sheets than through α helices.