pH-dependent Reduction Potential Research Articles

Theoretical Studies of Acetyl-CoA Synthase Catalytic Mechanism

DFT calculations were performed for the A-cluster from the enzyme Acetyl-CoA synthase (ACS). The acid constants (pKa), reduction potentials, and pH-dependent reduction potential for the A-cluster with different oxidation states and ligands were calculated. Good agreement of the reduction potentials, dependent on pH in the experiment, was obtained. On the basis of the calculations, a mechanism for the methylation reaction involving two–electron reduction and protonation on the proximal nickel atom of the reduced A-cluster is proposed.

Localized-Plasmon Voltammetry to Detect pH Dependent Gold Oxidation

Localized-plasmon voltammetry (LPV) bears great potential for electrochemical sensing applications beyond conventional cyclic voltammetry. In order to determine the limitations of this method, it is of utmost necessity to investigate the response toward chemical instability of the plasmonic electrode. We therefore investigated electrooxidation of a gold nanowire array with LPV in acidic electrolytes with different pH values. LPV shows excellent agreement with simultaneously recorded cyclic voltammograms up to the onset of oxygen evolution. Beyond that point, LPV still appears to provide meaningful signals. Further, with LPV the pH dependent reduction potentials of electrochemically grown gold oxides were determined and show a linear characteristic over the investigated pH range according to Nernst’s equation.

Using Hyperfine Electron Paramagnetic Resonance Spectroscopy to Define the Proton-Coupled Electron Transfer Reaction at Fe-S Cluster N2 in Respiratory Complex I.

Energy-transducing respiratory complex I (NADH:ubiquinone oxidoreductase) is one of the largest and most complicated enzymes in mammalian cells. Here, we used hyperfine electron paramagnetic resonance (EPR) spectroscopic methods, combined with site-directed mutagenesis, to determine the mechanism of a single proton-coupled electron transfer reaction at one of eight iron-sulfur clusters in complex I, [4Fe-4S] cluster N2. N2 is the terminal cluster of the enzyme's intramolecular electron-transfer chain and the electron donor to ubiquinone. Because of its position and pH-dependent reduction potential, N2 has long been considered a candidate for the elusive "energy-coupling" site in complex I at which energy generated by the redox reaction is used to initiate proton translocation. Here, we used hyperfine sublevel correlation (HYSCORE) spectroscopy, including relaxation-filtered hyperfine and single-matched resonance transfer (SMART) HYSCORE, to detect two weakly coupled exchangeable protons near N2. We assign the larger coupling with A(1H) = [-3.0, -3.0, 8.7] MHz to the exchangeable proton of a conserved histidine and conclude that the histidine is hydrogen-bonded to N2, tuning its reduction potential. The histidine protonation state responds to the cluster oxidation state, but the two are not coupled sufficiently strongly to catalyze a stoichiometric and efficient energy transduction reaction. We thus exclude cluster N2, despite its proton-coupled electron transfer chemistry, as the energy-coupling site in complex I. Our work demonstrates the capability of pulse EPR methods for providing detailed information on the properties of individual protons in even the most challenging of energy-converting enzymes.

A Flavin-Based pH Flow Battery That Recharges with Waste Heat or Carbon Dioxide Emissions

Currently, two enormous potential energy sources remain largely untapped: waste heat from energy production and industrial processes and carbon dioxide emissions from power plants. Industrial sites and nuclear plants produce enough potential energy in the form of waste heat to meet approximately half the U.S. energy demand (29000 TWh in 2013), and CO2 emissions can theoretically provide over one-third of the electricity demand (4078 TWh in 2015). Current technologies used to capture this potential energy are inefficient and/or use expensive materials. Here we propose a novel pH flow battery, which uses pH-dependent redox couples to extract energy from a pH gradient created from either low-grade waste heat or CO2 flue gases. For waste heat, ammonia (NH3) is dissolved in water to create a pH gradient that can be reversed by distillation using low-grade waste heat. Similarly, CO2 dissolved in water forms carbonic acid (H2CO3), which dissociates into bicarbonate (HCO3 -) and protons (H+) to create an acidic solution that can be subsequently removed by sparging with air. The goal of the study is to test the viability of the pH flow battery compared to other technologies. Cyclic voltammetry was used to test a suite of compounds to determine the pH dependency of reduction potentials, electrode kinetics, and compound stability. For sparingly soluble compounds, we evaluated the use of a hydrotropic agent, nicotinamide (vitamin B3), to increase solubility. Ideal compounds were then tested in a pH flow battery to evaluate power production, the ability to recharge the battery with waste heat or CO2, and cycling stability. From the compound survey, several Flavin-based compounds were identified as candidates due to fast electrode kinetics, pH-dependent reduction potentials over a wide pH range, and stability in oxygen. Nicotinamide added in high concentrations was shown to increase solubility of Flavin compounds at low pH and promising results were obtained from preliminary tests of the pH flow battery.

Reactive sites and course of reduction in the Rieske protein.

Rieske proteins play an essential role in electron transfer in the bc 1 complex. Rieske proteins contain a [2Fe-2S] cluster with one iron ligated by two histidines and the other iron ligated by two cysteines. All Rieske proteins have pH-dependent reduction potentials with the histidines ligating the cluster deprotonating in response to increases in pH. The addition of diethylpyrocarbonate (DEPC) modifies deprotonated histidines. The previous studies on the isolated Thermus thermophilus Rieske protein have used large excesses of DEPC, and this study examines what amino acids become modified under different molar equivalents of DEPC to protein. Increasing amounts of DEPC result in more modification, and higher pH values result in faster reaction. Upon modification, the protein also becomes reduced and ~6 equivalents of DEPC are needed for 50% of the reduction to occur. Which amino acids are modified first also points to the most reactive species on the protein. Mass spectrometry analysis shows that lysine 68 is the most reactive amino acid, followed by the ligating histidine 154 and two other surfaces lysines, 76 and 43. The modification of the ligating histidine at low numbers of DEPC equivalents and correlation with a similar number of equivalents needed to reduce the protein shows that this histidine can interact with neighboring groups, and these results can be extended to the protein within the bc 1 complex, where interaction with neighboring residues or molecules may allow reduction to occur. These results may shed light on how Rieske transfers electrons and protons in the bc 1 complex.

Coupling of Protonation, Reduction, and Conformational Change in azurin from Pseudomonas aeruginosa Investigated with Free Energy Measures of Cooperativity

We used free energy calculations within a continuum electrostatics model to analyze the coupling of protonation, reduction, and conformational change in azurin from Pseudomonas aeruginosa (PaAz). PaAz was characterized extensively with a variety of experimental methods. Experimentally determined pK(a) values and pH-dependent reduction potentials are used to validate our computational model. It is well-known from experiment that the reduction of the copper center is coupled to the protonation of at least two titratable residues (His-35 and His-83) and to the flip of the peptide bond between Pro-36 and Gly-37. Free energy measures of cooperativity are used for a detailed analysis of the coupling between protonation, reduction, and conformational change in PaAz. The reduction of the copper center, the protonation of His-35, and peptide flip are shown to be cooperative. Our results show that cooperativity free energies are useful in detecting and quantifying thermodynamic coupling between events in biomolecular systems. The protonation of His-35 and the peptide flip are found to be so tightly coupled that these events happen effectively concerted. This concerted change results in a marked alteration of the electrostatic surface potential of azurin that might affect the interaction of azurin with its binding partners.

Examination of n = 2 reaction mechanisms that reproduce pH-dependent reduction potentials

Flavo- and quinoproteins often exhibit trends in reduction potentials that approximate −30 mV per pH unit. At least five different reaction mechanisms can model the behavior described by the following Nernst equation: E observed = E 0 ′ + ( 0.059 / 2 ) log [ H + ] . The pH-reduction potential profile described by this equation can be reproduced by various models using different reaction mechanisms, and these mechanisms yield different reduction potentials and acid dissociation constants. In order to understand these discrepancies, this article discusses how the various methods reproduce this pH-dependent Nernst equation.

Voltammetric simulations of multiple electron transfer/proton transfer coupled reactions: flavin adenine dinucleotide as a model system

Cyclic voltammograms were simulated using DigiSim software for reaction mechanisms involving multiple electron transfer steps coupled to proton transfer. Specifically, the overall reaction mechanism of the form O + 2e − + 2H + ⇌ RH 2 was used to simulate experimental reduction potentials as a function of pH. The pH-dependent reduction potentials reported in the literature for flavodoxin and free flavin adenine dinucleotide were simulated based on selected reduction potentials and acid dissociation constants. Relationships between reduction potentials and acid dissociation constants are presented to model n=1 and n=2 reaction mechanisms and one- and two-proton-coupled redox reactions. Experimental parameters used in the simulations were selected such that the electron and proton transfer reactions were not rate limiting, and therefore these simulated reactions involve thermodynamic coupling rather than concerted kinetic processes.

Reduction potentials of Rieske clusters: importance of the coupling between oxidation state and histidine protonation state.

Rieske [2Fe-2S] clusters can be classified into two groups, depending on their reduction potentials. Typical high-potential Rieske proteins have pH-dependent reduction potentials between +350 and +150 mV at pH 7, and low-potential Rieske proteins have pH-independent potentials of around -150 mV at pH 7. The pH dependence of the former group is attributed to coupled deprotonation of the two histidine ligands. Protein-film voltammetry has been used to compare three Rieske proteins: the high-potential Rieske proteins from Rhodobacter sphaeroides (RsRp) and Thermus thermophilus (TtRp) and the low-potential Rieske ferredoxin from Burkholderia sp. strain LB400 (BphF). RsRp and TtRp differ because there is a cluster to serine hydrogen bond in RsRp, which raises its potential by 140 mV. BphF lacks five hydrogen bonds to the cluster and an adjacent disulfide bond. Voltammetry measurements between pH 3 and 14 reveal that all the proteins, including BphF, have pH-dependent reduction potentials with remarkably similar overall profiles. Relative to RsRp and TtRp, the potential versus pH curve of BphF is shifted to lower potential and higher pH, and the pK(a) values of the histidine ligands of the oxidized and reduced cluster are closer together. Therefore, in addition to simple electrostatic effects on E and pK(a), the reduction potentials of Rieske clusters are determined by the degree of coupling between cluster oxidation state and histidine protonation state. Implications for the mechanism of quinol oxidation at the Q(O) site of the cytochrome bc(1) and b(6)f complexes are discussed.

Terpsichorean movements of pentaammineruthenium on pyrimidine and isocytosine ligands.

Pentaammineruthenium moves on ambidentate nitrogen heterocycles by both rotation and linkage isomerization, which may affect the biological activity of potential ruthenium metallopharmaceuticals. The rapid rotation rates of [(NH3)5RuIII] coordinated to the exocyclic nitrogens of isocytosine (ICyt) and 6-methylisocytosine (6MeICyt) have been determined by 1H NMR. Since these rotamers can be stabilized by hydrogen bonding between the coordinated ammines and the N1 and N3 endocyclic nitrogens, rotamerization is under pH control. Spectrophotometrically (UV-vis) measured pKa values for the two endocyclic sites for the ICyt complex are 2.78 and 9.98, and for 6MeICyt are 3.06 and 10.21, which are probably weighted averages for ionization from N3 and N1, respectively. Activation parameters for the rotamerizations were determined by variable-temperature NMR at pKa1 < pH < pKa2 for the complexes with (ICyt-kappa N2)-, (6MeICyt kappa N2)-, and 2AmPym kappa N2. For [(6MeICyt kappa N2)(-)-(NH3)5RuIII]2+, delta H* = 1.6 kcal/mol, delta S* = -37 cal/mol K, and Ea = 2.2 kcal/mol. Due to strong RuIII-N pi-bonding, the activation enthalpies are approximately 10 kcal lower than the expected values for the free ligands. Rotameric structure is correlated with pKa values, pH-dependent reduction potentials, and 1H NMR parameters. Linkage isomers of [(2AmPym)(NH3)5Ru]n+ are reported in which RuII is coordinated to the endocyclic nitrogen (N1) and RuIII to the exocyclic nitrogen (N2). The rate constant for the kappa N2-->kappa N1 isomerization as part of an ECE mechanism is 3.9 s-1 at pH 3. The pH dependence of the acid-catalyzed hydrolysis of [(2AmPym kappa N1)(NH3)5Ru]2+ is determined.

Redox properties of flavocytochrome c3 from Shewanella frigidimarina NCIMB400.

The thermodynamic and catalytic properties of flavocytochrome c3 from Shewanella frigidimarina have been studied using a combination of protein film voltammetry and solution methods. As measured by solution kinetics, maximum catalytic efficiencies for fumarate reduction (kcat/Km = 2.1 x 10(7) M-1 s-1 at pH 7.2) and succinate oxidation (kcat/Km = 933 M-1 s-1 at pH 8.5) confirm that flavocytochrome c3 is a unidirectional fumarate reductase. Very similar catalytic properties are observed for the enzyme adsorbed to monolayer coverage at a pyrolytic graphite "edge" electrode, thus confirming the validity of the electrochemical method for providing complementary information. In the absence of fumarate, the adsorbed enzyme displays a complex envelope of reversible redox signals which can be deconvoluted to yield the contributions from each active site. Importantly, the envelope is dominated by the two-electron signal due to FAD [E degrees ' = -152 mV vs the standard hydrogen electrode (SHE) at pH 7.0 and 24 degrees C] which enables quantitative examination of this center, the visible spectrum of which is otherwise masked by the intense absorption bands due to the hemes. The FAD behaves as a cooperative two-electron center with a pH-dependent reduction potential that is modulated (pKox at 6.5) by ionization of a nearby residue. In conjunction with the kinetic pKa values determined for the forward and reverse reactions (7.4 and 8.6, respectively), a mechanism for fumarate reduction, incorporating His365 and an anionic form of reduced FAD, is proposed. The reduction potentials of the four heme groups, estimated by analysis of the underlying envelope, are -102, -146, -196, and -238 mV versus the SHE at pH 7.0 and 24 degrees C and are comparable to those determined by redox potentiometry.

Novel Redox Chemistry of [3Fe−4S] Clusters: Electrochemical Characterization of the All-Fe(II) Form of the [3Fe−4S] Cluster Generated Reversibly in Various Proteins and Its Spectroscopic Investigation in Sulfolobus acidocaldarius Ferredoxin

The novel “hyper-reduced” form of protein-bound [3Fe−4S] clusters, which is two electron equivalents below the normal reduced form [3Fe−4S]0 and thus formally composed entirely of Fe(II) subsites, has been characterized by electrochemistry and by EPR, MCD, and UV/visible spectroscopy. The two-electron reduction of [3Fe−4S]0 has been studied for a range of proteins, in particular the 7Fe ferredoxins from Sulfolobus acidocaldarius, Desulfovibrio africanus, and Azotobacter vinelandii. In each case, the reaction is chemically reversible, the product is surprisingly inert, and the pH-dependent reduction potential is in the region of −700 mV vs SHE at pH 7, regardless of the identity of the protein. Protein film voltammetry of three different ferredoxins investigated in detail over a wide pH range shows that the novel species denoted as [3Fe−4S]2- is formed by a cooperative two-electron reduction of [3Fe−4S]0 and there is a net uptake of three protons relative to the all-Fe(III) state [3Fe−4S]1+. The protons ar...

Azotobacter vinelandii ferredoxin I. Aspartate 15 facilitates proton transfer to the reduced [3Fe-4S] cluster.

The [3Fe-4S]+/0 cluster of Azotobacter vinelandii ferredoxin I (AvFdI) has an unusually low and strongly pH-dependent reduction potential (E'0). The reduced cluster exists in two forms, depending upon pH, that exhibit substantially different magnetic circular dichroism (MCD) spectra. Recent studies have established that the MCD changes observed on decreasing the pH from 8.3 (alkaline form) to 6.0 (acid form) cannot be explained either by a change in spin state of the cluster (Stephens, P.J., Jensen, G.M., Devlin, F.J., Morgan, T.V., Stout, C. D., Martin, A.E., and Burgess, B.K. (1991) Biochemistry 30, 3200-3209) or by a major structural change (e.g. ligand exchange) (Stout, C.D. (1993) J. Biol. Chem. 268, 25920-25927). Here, we have examined the influence of aspartate 15 on the pH dependence of the spectroscopic and electrochemical properties of AvFdI by construction of a D15N mutant. Aspartate 15, which is salt-bridged to lysine 84 at the protein surface, is the closest ionizable residue to the [3Fe-4S] cluster. The results show that replacement of aspartate by asparagine results in an approximately 20-mV increase in E'0 for the [3Fe-4S]+/0 cluster at high pH concomitant with an approximately 0.8-pH unit decrease in the pK of the reduced form. The major pH dependence of E'0 is preserved as is the effect observed by MCD. These data eliminate the possibility that the MCD change is due to the presence of Asp-15 and support the conclusion that it originates in direct protonation of the [3Fe-4S]0 cluster, probably on a sulfide ion. Voltammetric studies show that interconversion between [3Fe-4S]+ and [3Fe-4S]0 at acidic pH involves rapid electron transfer followed by proton transfer (for reduction) and then proton transfer followed by electron transfer (for oxidation). Ionized aspartate 15 facilitates proton transfer. Thus, protonation and deprotonation are much slower for D15N relative to the native protein at pH > 5.5. Proton transfer reactions necessary for further reduction of the [3Fe-4S]0 cluster to the [3Fe-4S]- and [3Fe-4S]2- states are also retarded in D15N. The results suggest that the carboxylate-ammonium salt bridge afforded by Asp-15-Lys-84 conducts protons between the cluster and solvent H2O molecules. Overproduction of D15N FdI, but not native FdI, in A. vinelandii has a negative effect on the growth rate of the organism, suggesting that the rate of protonation or deprotonation of the [3Fe-4S]0 cluster may be important in vivo.

Electrochemical investigation of acid—base properties of ordered liquid systems

Acid—base properties of ordered media were investigated via potentiometry, polarography and electrochemical probes. Electrochemical probes have a pH-dependent reduction potential and their oxidized and reduced forms have a different affinity for aqueous and organic phases. Solutions of anionic, cationic and nonionic surfactants were investigated. One anionic and one cationic surfactant stabilized emulsion were studied. A water—dodecane—pentanol—anionic surfactant microemulsion and a water—heptane—butanol—cationic surfactant were also investigated for several compositions. In micellar solutions and emulsions, it was possible to standardize and use the classical glass electrode for pH values in the range 1–12. The hydrogen electrode was required in the microemulsion systems. The reduction of electrochemical probes was studied by polarography. It is shown that in the ordered media studied, the aqueous phase played the most important role in micellar solutions and in O/W emulsions, as far as acid—base properties were concerned. In microemulsions, the acid—base properties of the aqueous phase were very different to those of water. The alizarin probe could be reduced at a “local” pH of about 12 when the aqueous phase pH was only 6.

A totally synthetic histidine-2 ferredoxin: thermal stability and redox properties.

The entire polypeptide of Clostridium pasteurianum ferredoxin (Fd) with a site-substituted tyrosine-2----histidine-2 was synthesized using standard t-Boc procedures, reconstituted to the 2[4Fe-4S] holoprotein, and compared to synthetic C. pasteurianum and native Fds. Although histidine-2 is commonly found in thermostable clostridial Fds, the histidine-2 substitution into synthetic C. pasteurianum Fd did not significantly increase its thermostability. The reduction potential of synthetic histidine-2 Fd was -343 and -394 mV at pH 6.4 and 8.7, respectively, versus standard hydrogen electrode. Similarly, Clostridium thermosaccharolyticum Fd which naturally contains histidine-2 was previously determined to have a pH-dependent reduction potential [Smith, E.T., & Feinberg, B.A. (1990) J. Biol. Chem. 265, 14371-14376]. An electrostatic model was used to calculate the observed change in reduction potential with pH for a homologous ferredoxin with a known X-ray crystal structure containing a hypothetical histidine-2. In contrast, the reduction potential of both native C. pasteurianum Fd and synthetic Fd with the C. pasteurianum sequence was -400 mV versus standard hydrogen electrode and was pH-independent [Smith, E.T., Feinberg, B.A., Richards, J.H., & Tomich, J.M. (1991) J. Am. Chem. Soc. 113, 688-689]. On the basis of the above results, we conclude that the observed pH-dependent reduction potential for both synthetic and native ferredoxins that contain histidine-2 is attributable to the electrostatic interaction between histidine-2 and iron-sulfur cluster II which is approximately 6 A away.

Site-directed mutagenesis of Azotobacter vinelandii ferredoxin I. Changes in [4Fe-4S] cluster reduction potential and reactivity.

We have used site-directed mutagenesis to obtain two variants of Azotobacter vinelandii ferredoxin I (AvFdI), whose x-ray structures are now available. In the C20A protein, a ligand to the [4Fe-4S] cluster was removed whereas in the C24A mutant a free cysteine next to that cluster was removed. Like native FdI, both mutants contain one [4Fe-4S] cluster and one [3Fe-4S] cluster. The structure of C24A is very similar to that of native FdI, while the structure of C20A is rearranged in the region of the [4Fe-4S] cluster to allow it to use the free Cys-24 as a replacement ligand. Here we compare the properties of the native, C20A, and C24A proteins. Although all three proteins are O2 stable in vitro, the C20A protein is much less stable toward proteolysis than the other two in vivo. Spectroscopic results show that all three proteins exhibit the same general redox behavior during O2-oxidation and dithionite reduction. Electrochemical data show that the [3Fe-4S] clusters in all three proteins have the same pH-dependent reduction potentials (-425 mV versus SHE, pH 7.8), whereas the [4Fe-4S] cluster potentials vary over a approximately 150 mV range from -600 mV (C24A) to -647 mV (native) to -746 mV (C20A). Despite this variation in potential both the C20A and C24A proteins appear to be functional in vivo. Native FdI reacts with three equivalents of Fe(CN)3-(6) to form a paramagnetic species previously proposed to be a cysteinyl-disulfide radical. Neither the C20A nor the C24A variant undergoes this reaction, strongly suggesting that it involves the free Cys-24.

Redox-linked proton translocation in cytochrome oxidase: the importance of gating electron flow. The effects of slip in a model transducer

In at least one component of the mitochondrial respiratory chain, cytochrome c oxidase, exothermic electron transfer reactions are used to drive vectorial proton transport against an electrochemical hydrogen ion gradient across the mitochondrial inner membrane. The role of the gating of electrons (the regulation of the rates of electron transfer into and out of the proton transport site) in this coupling between electron transfer and proton pumping has been explored. The approach involves the solution of the steady-state rate equations pertinent to proton pump models which include, to various degrees, the uncoupled (i.e., not linked to proton pumping) electron transfer processes which are likely to occur in any real electron transfer-driven proton pump. This analysis furnishes a quantitative framework for examining the effects of variations in proton binding site pKas and metal center reduction potentials, the relationship between energy conservation efficiency and turnover rate, the conditions for maximum power output or minimum heat production, and required efficiency of the gating of electrons. Some novel conclusions emerge from the analysis, including: An efficient electron transfer-driven proton pump need not exhibit a pH-dependent reduction potential; Very efficient gating of electrons is required for efficient electron transfer driven proton pumping, especially when a reasonable correlation of electron transfer rate and electron transfer exoergonicity is assumed; and A consideration of the importance and possible mechanisms of the gating of electrons suggests that efficient proton pumping by CuA in cytochrome oxidase could, in principle, take place with structural changes confined to the immediate vicinity of the copper ion, while proton pumping by Fea would probably require conformational coupling between the iron and more remote structures in the enzyme. The conclusions are discussed with reference to proton pumping by cytochrome c oxidase, and some possible implications for oxidative phosphorylation are noted.

Heterogeneous catalysis and electrosorption phenomena in the cathodic reduction of acetylpyridines: Part II. Electrode kinetic investigations

The mechanism of the cathodic reduction of three isomeric acetylpyridines has been evaluated by electrode kinetic and preparative measurements at the DME and rotating rod electrodes of different solid metal electrode materials. The influence of electrosorbed molecules, which may either act as proton donors and asymmetric inductors (strychnine, camphoric acid) or are chemically inert surfactants (tetraalkylammonium ions) on the electrode kinetics, has been studied. Up to pH 10, acetylpyridines are reduced from their N-protonated form. If the strongly pH-dependent reduction potential of the ketone is close to the E pzc of the electrode, electrosorbed ketyl radicals are formed in a 1 e −/1 H + EC reaction sequence and the pinacol is formed by heterogeneous dimerization of these adsorbed radicals. This holds for the 2- and 4-acetylpyridine reduction at low pH (1<pH<3.5). At higher pH (pH>4) and in the absence of electrosorbed proton donors, the alcohol formation proceeds by eventual irreversible protonation of the pyridyl-benzylic carbanion which is formed at more negative potentials in a fast and reversible (2 e −/H +)-reduction step. Reduction of 2- and 4-acetylpyridine in the presence of adsorbed proton donors (1<pH<7) is speeded up due to the participation of the adsorbed proton donor in the initial 2 e −/H + equilibrium. Heterogeneous protonation of the ketyl radical brought about by adsorbed chiral proton donors (inductor acid) is one of the key steps in cathodic asymmetric electro-organic synthesis. The ketyl radicals formed by a 1 e −/1 H + protonation reduction step from the three ketones possess relatively different basicities: for 2-acetylpyridine the radical is a 10 7 times stronger base, and for 4-acetylpyridine a 10 5 times stronger base than for 3-acetylpyridine. For the least basic ketyl radical of 3-acetylpyridine, only surfactants which are stronger acids, such as camphoric acid, show a decrease in lifetime and a speeding up of electrode kinetics which yields an optical induction.