pH Dependence of charge transfer between tryptophan and tyrosine in dipeptides

Time-resolved absorption spectroscopy has been employed to study the directionality and rate of charge transfer in W-Y and Ac-W-Y dipeptides as a function of pH. Excitation with 266-nm nanosecond laser pulses produces both W (or [WH](+), depending on pH) and Y. Between pH 6 and 10, W to was found to oxidize Y with k(X)=9.0x10(4) s(-1) and 1.8x10(4) s(-1) for the W-Y and Ac-W-Y dipeptide systems, respectively. The intramolecular charge transfer rate increases as the pH is lowered over the range 6>pH>2. For 10W-Y(-) (Y(-), tyrosinate anion), with a rate constant of k(X)=1.2x10(5) s(-1). The dependence of charge transfer directionality between W and Y on pH is important to the enzymatic function of several model and natural biological systems as discussed here for ribonucleotide reductase.     

A comparative study of complex formation in the reactions of gold(III) with Gly-Gly, Gly-l-Ala and Gly-l-His dipeptides

Proton NMR spectroscopy was applied to study the reactions of the dipeptides glycyl-glycine (Gly-Gly) and glycyl-l-alanine (Gly-l-Ala) with hydrogen tetrachloridoaurate(III) (H[AuCl₄]). All reactions were performed at pH 2.0 and 3.0 and at 40 °C. The final products in these reactions were [Au(Gly-Gly-κ³N_G1,N_G2,O_G2)Cl] and [Au(Gly-l-Ala-κ³N_G,N_A,O_A)Cl] complexes. Tridentate coordination of the corresponding dipeptides and square-planar geometry of these Au(III) complexes was confirmed by NMR (¹H and ¹³C) spectroscopy. This study showed that at pH < 3.0 the Au(III) ion was able to deprotonate the amide nitrogen atom. However this displacement reaction was very slow and the total concentration of the corresponding Au(III)-peptide complex formed after 5 days was less than 60% for the Gly-l-Ala or 70% for the Gly-Gly dipeptide. The kinetic data of the reactions between the Gly-Gly and Gly-l-Ala dipeptides and [AuCl₄]⁻ were compared with those for the histidine-containing Gly-l-His dipeptide. The differences in the reactivity of these three dipeptides with the Au(III) ion are discussed.     

Effective heterogeneous hydrolysis of phosphodiester by pyridine-containing metallopolymers

The copper (II) complex of a simple pyridine- and amide-containing copolymer serves as an effective catalyst for heterogeneous hydrolysis of the prototypical phosphodiester substrate bis(p-nitrophenyl)phosphate at pH 8.0 and 25 °C. The catalysis has a first-order rate constant of k_cat = 8.3 × 10⁻⁶ s⁻¹, corresponding to a catalytic proficiency of 75-thousand folds relative to the uncatalyzed hydrolysis with a rate constant of k₀ = 1.1 × 10⁻¹⁰ s⁻¹ in aqueous buffer solution at pH 8.0. This observation suggests that polymers can be designed to include various functional groups feasible for effective metal-centered catalysis of phosphodiester hydrolysis.     

Aqueous solution chemistry of dichloro[S-methylcysteine(N,S)]platinum(II) isomers and their hydrolysis products

Rate and equilibrium constants at 25 °C, pH ∼ 1, and ionic strength 0.10 for hydrolysis of the two non-equivalent chlorides of dichloro[S-methyl-l-cysteine(N,S)]platinum(II) isomers, denoted [PtCl₂(SmecysH)], and the resultant chloro-aqua species have been determined by NMR, potentiometric, and spectrophotometric methods. Though hydrolysis constants, K_h, for the two chlorides are similar (pK_h = 4-5), the rate of hydrolysis of the chloride trans to coordinated S, k_h = 3.4 × 10⁻³ s⁻¹, is 2-3 orders of magnitude faster than the k_h for the other chloride, 2.3 × 10⁻⁶ s⁻¹, and for the cancer drug cisplatin, cis-[PtCl₂(NH₃)₂], 5.2 × 10⁻⁵ s⁻¹. Relative rates of hydrolysis determined under three different experimental conditions (pH ∼ 1 in 0.10 M HNO₃, high pH in 0.10 M NaOH, and at low pH with Ag⁺ assistance) are consistent: the Cl trans to S is 100-1000 times more labile than the Cl cis to S. Potentiometric and NMR methods were also used to estimate pK_a values of all aqua species, which are comparable to values reported for corresponding aqua species derived from cisplatin.     

Downstream reaction of cisplatin with methionine-containing peptides: pH-dependent competition between hydrolytic cleavage and macrochelation

The pH- and time-dependent reactions of the antitumor drug cisplatin, cis-[PtCl₂(NH₃)₂], with the methionine-containing peptides Ac-Met-Gly-OH, Ac-Met-Pro-OH, Ac-Met-Pro-Gly-Gly-OH and Ac-Gly-Met-Pro-Gly-Gly-OH (Gly = glycyl, Met = d-methionyl, Pro = L-prolyl) at 313 K have been investigated by high performance liquid chromatography, mass spectrometry and nuclear magnetic resonance. As a result of the strong trans influence of the methionyl S_M atom, initial Pt-S_M binding at pH > 5 is followed by a rapid formation of tridentate machrochelates for the N-acetylated peptides. The site trans to S_M is occupied by a carboxylate O atom in the case of the κ³S_M,N_M,O_G/P macrochelates of the dipeptides and by the C-terminal glycylamide N_G2 atom for the κ³S_M,O_M,N_G2 macrochelate of Ac-Met-Pro-Gly-Gly-OH. Cisplatin simultaneously mediates the rapid hydrolytic cleavage of the Met-X (X = Gly, Pro) amide bond for both dipeptides over the whole range 2.8 ? pH ? 10.0. The released amino acids X react with the resulting κ²S_M, N_M chelate of N-acetylmethionine to afford mixed κS_M:κ²N_x,O_x complexes of the type cis-[Pt(NH₃)(Ac-Met-OH-κS)(H-X-O-κ²N_x,O_x)]⁺ as final products at pH < 5 for X = Gly and pH < 8 for X = Pro. In contrast to the dipeptides, hydrolytic cleavage of the Met-Pro amide bond in Ac-Met-Pro-Gly-Gly-OH at pH > 5 is significantly inhibited by the presence of high concentrations of the macrochelate [Pt(NH₃)(Ac-Met-Pro-Gly-Gly)-κ³S_M,O_M,N_G2]⁺. Downstream hydrolysis of the Met-Gly amide bond is competitive with upstream Ac-Gly cleavage for Ac-Gly-Met-Pro-Gly-Gly-OH at pH < 4.5.     

Optical properties of Ag(tripod)X with tripod = 1,1,1-tris(diphenyl-phosphinomethyl)ethane and X = Cl and I: Intraligand and ligand-to-ligand charge transfer

The complexes Ag^I(tripod)X with tripod = 1,1,1-tris(diphenylphosphinomethyl)ethane and X⁻ = Cl⁻ and I⁻ are luminescent in solution at r.t. It is suggested that the emission is a phosphorescence which originates from a tripod intraligand state for X = Cl (λ_max = 464 nm) and a X⁻ → tripod ligand-to-ligand charge transfer state for X = I (λ_max = 482 nm).     

Kinetic and thermodynamic properties of the folding and assembly of formate dehydrogenase

The folding mechanism and stability of dimeric formate dehydrogenase from Candida methylica was analysed by exposure to denaturing agents and to heat. Equilibrium denaturation data yielded a dissociation constant of about 10⁻¹³ M for assembly of the protein from unfolded chains and the kinetics of refolding and unfolding revealed that the overall process comprises two steps. In the first step a marginally stable folded monomeric state is formed at a rate (k₁) of about 2 × 10⁻³ s⁻¹ (by deduction k₋₁ is about10⁻⁴ s⁻¹) and assembles into the active dimeric state with a bimolecular rate constant (k₂) of about 2 × 10⁴ M⁻¹ s⁻¹. The rate of dissociation of the dimeric state in physiological conditions is extremely slow (k₋₂ ∼ 3 × 10⁻⁷ s⁻¹).     

Synthesis, characterization and antioxidant activity of water soluble Mn complexes of sulphonato-substituted Schiff base ligands

Two new Mn^III complexes Na[Mn(5-SO₃-salpnOH)(H₂O)] ⋅ 5H₂O (1) and Na[Mn(5-SO₃-salpn)(MeOH)] ⋅ 4H₂O (2) (5-SO₃-salpnOH = 1,3-bis(5-sulphonatosalicylidenamino)propan-2-ol, 5-SO₃-salpn = 1,3-bis(5-sulphonatosalicylidenamino)propane) have been prepared and characterized. Electrospray ionization-mass spectrometry, UV-visible and ¹H NMR spectroscopic studies showed that the two complexes exist in solution as monoanions [Mn(5-SO₃-salpn(OH))(solvent)₂]⁻, with the ligand bound to Mn^III through the two phenolato-O and two imino-N atoms located in the equatorial plane. The E_1/2 of the Mn^III/Mn^II couple (−47.11 (1) and −77.80 mV (2) vs. Ag/AgCl) allows these complexes to efficiently catalyze the dismutation of , with catalytic rate constants 2.4 × 10⁶ (1) and 3.6 × 10⁶ (2) M⁻¹ s⁻¹, and IC₅₀ values of 1.14 (1) and 0.77 (2) μM, obtained through the nitro blue tetrazolium photoreduction inhibition superoxide dismutase assay, in aqueous solution of pH 7.8. The two complexes are also able to disproportionate up to 250 equivalents of H₂O₂ in aqueous solution of pH 8.0, with initial turnover rates of 178 (1) and 25.2 (2) mM H₂O₂ min⁻¹ mM⁻¹ catalyst⁻¹. Their dual superoxide dismutase/catalase activity renders these compounds particularly attractive as catalytic antioxidants.     

Acrylonitrile Quenching of Trp Phosphorescence in Proteins: A Probe of the Internal Flexibility of the Globular Fold

Quenching of Trp phosphorescence in proteins by diffusion of solutes of various molecular sizes unveils the frequency-amplitude of structural fluctuations. To cover the sizes gap between O₂ and acrylamide, we examined the potential of acrylonitrile to probe conformational flexibility of proteins. The distance dependence of the through-space acrylonitrile quenching rate was determined in a glass at 77 K, with the indole analog 2-(3-indoyl) ethyl phenyl ketone. Intensity and decay kinetics data were fitted to a rate, k(r) = k₀ exp[−(r − r₀)/r_e], with an attenuation length r_e = 0.03 nm and a contact rate k₀ = 3.6 × 10¹⁰ s⁻¹. At ambient temperature, the bimolecular quenching rate constant (kq) was determined for a series of proteins, appositely selected to test the importance of factors such as the degree of Trp burial and structural rigidity. Relative to kq = 1.9 × 10⁹ M⁻¹s⁻¹ for free Trp in water, in proteins kq ranged from 6.5 × 10⁶ M⁻¹s⁻¹ for superficial sites to 1.3 × 10² M⁻¹s⁻¹ for deep cores. The short-range nature of the interaction and the direct correlation between kq and structural flexibility attest that in the microsecond-second timescale of phosphorescence acrylonitrile readily penetrates even compact protein cores and exhibits significant sensitivity to variations in dynamical structure of the globular fold.     

Kinetic studies of the reaction of heme-thiolate enzyme chloroperoxidase with peroxynitrite

The kinetics of the reaction of chloroperoxidase with peroxynitrite was studied under neutral and acidic pH by stopped-flow spectrophotometry. Chloroperoxidase catalyzed peroxynitrite decay with the rate constant, k_c, increasing with decreasing pH. The values of k_c obtained at pH 5.1, 6.1 and 7.1 were equal to: (1.96 ± 0.03) × 10⁶, (1.63 ± 0.04) × 10⁶ and (0.71 ± 0.01) × 10⁶ M⁻¹ s⁻¹, respectively. Chloroperoxidase was converted to compound II by peroxynitrite with pH-dependent rate constants: (12.3 ± 0.4) × 10⁶ and (3.8 ± 0.3) × 10⁶ M⁻¹ s⁻¹ at pH 5.1 and 7.1, respectively. After most of peroxynitrite had disappeared, the conversion of compound II into the ferric form of chloroperoxidase was observed. The recovery of the native enzyme was completed within 1 s and 5 s at pH 5.1 and 7.1, respectively. The possible reaction mechanisms of the catalytic decomposition of peroxynitrite by chloroperoxidase are discussed.     

Characterization of the adenine nucleoside specific phosphorylase of Bacillus cereus

Adenosine phosphorylase, a purine nucleoside phosphorylase endowed with high specificity for adenine nucleosides, was purified 117-fold from vegetative forms of Bacillus cereus. The purification procedure included ammonium sulphate fractionation, pH 4 treatment, ion exchange chromatography on DEAE-Sephacel, gel filtration on Sephacryl S-300 HR and affinity chromatography on N⁶-adenosyl agarose. The enzyme shows a good stability to both temperature and pH. It appears to be a homohexamer of 164 ± 5 kDa. Kinetic characterization confirmed the specificity of this phosphorylase for 6-aminopurine nucleosides. Adenosine was the preferred substrate for nucleoside phosphorolysis (k_cat/K_m 2.1 × 10⁶ s^− 1 M^− 1), followed by 2′-deoxyadenosine (k_cat/K_m 4.2 × 10⁵ s^− 1 M^− 1). Apparently, the low specificity of adenosine phosphorylase towards 6-oxopurine nucleosides is due to a slow catalytic rate rather than to poor substrate binding.     

Photoinduced electron transfer and energy transfer reactions of hydroxo-(2,2′:6′,2″-terpyridine) platinum(II)

The luminescent complex [Pt(terpy)OH]BF₄ undergoes photoinduced electron transfer reactions with phenyl amine electron donors and nitrophenyl electron acceptors. Stern-Volmer analysis of the quenching of metal-to-ligand charge transfer phosphorescence (³MLCT) was used to calculate bimolecular rate constants for electron transfer. Rate constants vary from 10⁸ to >10¹⁰ M⁻¹ s⁻¹, depending on the thermodynamic driving force of the electron transfer reaction, with rate constants indicating that [Pt(terpy)OH]BF₄* is a powerful photo-oxidant. Aromatic triplet energy acceptors can also quench the ³MLCT emission.     

Kinetics and mechanism of the reactions of ozone with superoxo, hydroperoxo, and hydrido complexes of rhodium(III)

Oxidation of the title complexes with ozone takes place by hydrogen atom, hydride, and electron transfer mechanisms. The reaction with (NH₃)₄(H₂O)RhH²⁺ is a two electron process, believed to involve hydride transfer with a rate constant k = (2.2 ± 0.2) × 10⁵ M⁻¹ s⁻¹ and an isotope effect k_H/k_D = 2. The oxidation of (NH₃)₄(H₂O)RhOOH²⁺ to (NH₃)₄(H₂O)RhOO²⁺ by an apparent hydrogen atom transfer is quantitative and fast, k = (6.9 ± 0.3) × 10³ M⁻¹ s⁻¹, and constitutes a useful route for the preparation of the superoxo complex. The latter is also oxidized by ozone, but more slowly, k = 480 ± 50 M⁻¹ s⁻¹.     

Large catalase based bioelectrode for biosensor application

A large catalase (CAT) (Mr ~ 90 kDa), immobilized on multiwalled carbon nanotubes—Nafion^® (MWCNT-NF) matrix and encapsulated with polyethylenimine (PEI) on glassy carbon electrode (GCE), showed a pair of nearly reversible cyclic voltammetric peaks for Fe^(III)/Fe^(II) couple with formal potential of about −0.45 V (vs. Ag/AgCl electrode at pH 7.5). PEI significantly reduced the charge transfer resistance and stabilized the bioelectrode through electrostatic interaction. The electron transfer rate constant and surface coverage of the immobilized CAT were 1.05 ± 0.2 s⁻¹ and 2.1 × 10⁻¹⁰ mol cm⁻², respectively. Studies on electrocatalytic activity and kinetics of GCE/MWCNT-NF/CAT/PEI for hydrogen peroxide (H₂O₂) showed the apparent Michaelis-Menten constant of 3 mM, linear response in the range of 10 μM to 5 mM, response time of ~ 2 s for steady state current, and detection limit of ~ 1 μM. A high operational and storage stability was also demonstrated for the bioelectrode. Hence, the direct electrochemistry of the large catalase and its potential biosensor application have been established through this investigation.     

Cyanide and chloride exchange on homoleptic gold(III) square-planar complexes: variable pressure kinetic investigation by heteronuclear NMR

Kinetic studies of X⁻ exchange on [AuX₄]⁻ square-planar complexes (where X=Cl⁻ and CN⁻) were performed at acidic pH in the case of chloride system and as a function of pH for the cyanide one. Chloride NMR study (330-365 K) gives a second-order rate law on [AuCl₄]⁻ with the kinetic parameters: (k₂^Au,Cl)²⁹⁸=0.56±0.03 s⁻¹ mol⁻¹ kg; ΔH₂^‡ Au,Cl=65.1±1 kJ mol⁻¹; ΔS₂^‡ Au,Cl=−31.3±3 J mol⁻¹ K⁻¹ and ΔV₂^‡ Au,Cl=−14±2 cm³ mol⁻¹. The variable pressure data clearly indicate the operation of an I_a or A mechanism for this exchange pathway. The proton exchange on HCN was determined by ¹³C NMR as a function of pH and the rate constant of the three reaction pathways involving H₂O, OH⁻ and CN⁻ were determined: k₀^HCN,H=113±17 s⁻¹, k₁^HCN,H=(2.9±0.7)×10⁹ s⁻¹ mol⁻¹ kg and k₂^HCN,H=(0.6±0.2)×10⁶ s⁻¹ mol⁻¹ kg at 298.1 K. The rate law of the cyanide exchange on [Au(CN)₄]⁻ was found to be second order with the following kinetic parameters: (k₂^Au,CN)²⁹⁸=6240±85 s⁻¹ mol⁻¹ kg, ΔH₂^‡ Au,CN=40.0±0.8 kJ mol⁻¹, ΔS₂^‡ Au,CN=−37.8±3 J mol⁻¹ K⁻¹ and ΔV₂^‡ Au,CN=+2±1 cm³ mol⁻¹. The rate constant observed varies about nine orders of magnitude depending on the pH and HCN does not act as a nucleophile. The observed rate constant of X⁻ exchange on [AuX₄]⁻ are two or three orders of magnitude faster than the Pt(II) analogue.     

Characterisation of the electron transfer and complex formation between Flavodoxin from D. vulgaris and the haem domain of Cytochrome P450 BM3 from B. megaterium

Investigation of the complex formation and electron transfer kinetics between P450 BMP and flavodoxin was carried out following the suggested involvement of flavodoxin in modulating the electron transfer to BMP in artificial redox chains bound to an electrode surface. While electron transfer measurements show the formation of a tightly bound complex, the NMR data indicate the formation of shortly lived complexes. The measured k_obs ranged from 24.2 s^− 1 to 44.1 s^− 1 with k_on ranging from 0.07 × 10⁶ to 1.1 × 10⁶ s^− 1M^− 1 and K_d ranging from 300 μM to 24 μM in buffers of different ionic strength. This apparent contradiction is due to the existence of two events in the complex formation prior to electron transfer. A stable complex is initially formed. Within such tightly bound complex, flavodoxin rocks rapidly between different positions. The rocking of the bound flavodoxin between several different orientations gives rise to the transient complexes in fast exchange as observed in the NMR experiments. Docking simulations with two different approaches support the theory that there is no highly specific orientation in the complex, but instead one side of the flavodoxin binds the P450 with high overall affinity but with a number of different orientations. The level of functionality of each orientation is dependent on the distance between cofactors, which can vary between 8 and 25 Å, with some of the transient complexes showing distances compatible with the measured electron transfer rate constants.     

Solvent exchange, solvent interchange, aquation and isomerisation reactions of cis- and trans-[Co(tmen)2(NCMe)2] in water, Me2SO and MeCN: kinetics and stereochemistry

The synthesis and characterisation of cis- and trans-[Co(tmen)₂(NCCH₃)₂](ClO₄)₃ are described. Solvolysis rates have been measured by both ¹H NMR spectroscopy and UV-Vis spectrophotometry in dimethyl sulfoxide at 298.2 K. The cis isomer undergoes solvolysis by consecutive first-order reactions, k₁=5.61 × 10⁻⁴ and k₂=5.35 × 10⁻⁴ s⁻¹, each with steric retention. The measured solvolysis rate (single step reaction) for the trans isomer is k=1.54 × 10⁻⁵ s⁻¹. The solvent exchange rates have been measured by ¹H NMR spectroscopy in CD₃CN at 298.2 K: k_ex(cis)=k_ct + k_cc=2.0 × 10⁻⁵ and k_ex(trans)=k_tc + k_tt=4.56 × 10⁻⁶ s⁻¹. From these data, the measured cis-trans isomerisation rate (1.71 × 10⁻⁶ s⁻¹) and equilibrium position in CH₃CN (17% trans), the steric course for substitution in the exchange processes has been determined: trans reactant - 69% trans product; cis reactant - 99% cis product. Aquation rates for cis- and trans-[Co(tmen)₂(NCCH₃)₂](ClO₄)₃ have also been determined spectrophotometrically and by NMR; k_cis=1.3 × 10⁻⁴ and k_trans=2.7 × 10⁻⁵ s⁻¹. In both cases the steric course for the primary aquation step is indeterminate because the subsequent steps are faster. Where data are available, the [Co(tmen)₂X₂]ⁿ⁺ complexes are found to be consistently much more reactive than their [Co(en)₂X₂]ⁿ⁺ analogues.     

Stabilization of charge separation and cardiolipin confinement in antenna-reaction center complexes purified from Rhodobacter sphaeroides

The reaction center-light harvesting complex 1 (RC-LH1) purified from the photosynthetic bacterium Rhodobacter sphaeroides has been studied with respect to the kinetics of charge recombination and to the phospholipid and ubiquinone (UQ) complements tightly associated with it. In the antenna-RC complexes, at 6.5 < pH < 9.0, P⁺Q_B⁻ recombines with a pH independent average rate constant <k> more than three times smaller than that measured in LH1-deprived RCs. At increasing pH values, for which <k> increases, the deceleration observed in RC-LH1 complexes is reduced, vanishing at pH > 11.0. In both systems kinetics are described by a continuous rate distribution, which broadens at pH > 9.5, revealing a strong kinetic heterogeneity, more pronounced in the RC-LH1 complex. In the presence of the antenna the Q_AQ_B⁻ state is stabilized by about 40 meV at 6.5 < pH < 9.0, while it is destabilized at pH > 11. The phospholipid/RC and UQ/RC ratios have been compared in chromatophore membranes, in RC-LH1 complexes and in the isolated peripheral antenna (LH2). The UQ concentration in the lipid phase of the RC-LH1 complexes is about one order of magnitude larger than the average concentration in chromatophores and in LH2 complexes. Following detergent washing RC-LH1 complexes retain 80-90 phospholipid and 10-15 ubiquinone molecules per monomer. The fractional composition of the lipid domain tightly bound to the RC-LH1 (determined by TLC and ³¹P-NMR) differs markedly from that of chromatophores and of the peripheral antenna. The content of cardiolipin, close to 10% weight in chromatophores and LH2 complexes, becomes dominant in the RC-LH1 complexes. We propose that the quinone and cardiolipin confinement observed in core complexes reflects the in vivo heterogeneous distributions of these components. Stabilization of the charge separated state in the RC-LH1 complexes is tentatively ascribed to local electrostatic perturbations due to cardiolipin.     

Comparative study of catalase-peroxidases (KatGs)

Catalase-peroxidases or KatGs from seven different organisms, including Archaeoglobus fulgidus,Bacillus stearothermophilus, Burkholderia pseudomallei, Escherichia coli, Mycobacterium tuberculosis, Rhodobacter capsulatus and Synechocystis PCC 6803, have been characterized to provide a comparative picture of their respective properties. Collectively, the enzymes exhibit similar turnover rates with the catalase and peroxidase reactions varying between 4900 and 15,900 s⁻¹ and 8-25 s⁻¹, respectively. The seven enzymes also exhibited similar pH optima for the peroxidase (4.25-5.0) and catalase reactions (5.75), and high sensitivity to azide and cyanide with IC₅₀ values of 0.2-20 μM and 50-170 μM, respectively. The K_Ms of the enzymes for H₂O₂ in the catalase reaction were relatively invariant between 3 and 5 mM at pH 7.0, but increased to values ranging from 20 to 225 mM at pH 5, consistent with protonation of the distal histidine (pKa approximately 6.2) interfering with H₂O₂ binding to Cpd I. The catalatic k_cat was 2- to 3-fold higher at pH 5 compared to pH 7, consistent with the uptake of a proton being involved in the reduction of Cpd I. The turnover rates for the INH lyase and isonicotinoyl-NAD synthase reactions, responsible for the activation of isoniazid as an anti-tubercular drug, were also similar across the seven enzymes, but considerably slower, at 0.5 and 0.002 s⁻¹, respectively. Only the NADH oxidase reaction varied more widely between 10⁻⁴ and 10⁻² s⁻¹ with the fastest rate being exhibited by the enzyme from B. pseudomallei.     

Kinetics and mechanism of the reduction of the molybdatopentaamminecobalt(III) ion by aqueous sulfite and aqueous potassium hexacyanoferrate(II)

A detailed investigation on the oxidation of aqueous sulfite and aqueous potassium hexacyanoferrate(II) by the title complex ion has been carried out using the stopped-flow technique over the ranges, 0.01≤[S(IV)]_T≤0.05 mol dm⁻³, 4.47≤pH≤5.12, and 24.9≤θ≤37.6 °C and at ionic strength 1.0 mol dm⁻³ (NaNO₃) for aqueous sulfite and 0.01≤[Fe(CN)₆ ⁴⁻]≤0.11 mol dm⁻³, 4.54≤pH≤5.63, and 25.0≤θ≤35.3 °C and at ionic strength 1.0 or 3.0 mol dm⁻³ (NaNO₃) for the hexacyanoferrate(II) ion. Both redox processes are dependent on pH and reductant concentration in a complex manner, that is, for the reaction with aqueous sulfite, k_obs={(k₁K₁K₂K₃+k₂K₁K₄[H⁺])[S(IV)]_T]/([H⁺]²+K₁[H⁺]+K₁K₂) and for the hexacyanoferrate(II) ion, k_obs={(k₁K₃K₄K₅+k₂K₃K₆[H⁺])[Fe(CN)₆ ⁴⁻]_T)/([H⁺]²+K₃[H⁺]+K₃K₄). At 25.0 °C, the value of k₁′ (the composite of k₁K₃) is 0.77±0.07 mol⁻¹ dm³ s⁻¹, while the value of k₂′ (the composite of k₂K₄) is (3.78±0.17)×10⁻² mol⁻¹ dm³ s⁻¹ for aqueous sulfite. For the hexacyanoferrate(II) ion, k₁′ (the composite of k₁K₅) is 1.13±0.01 mol⁻¹ dm³ s⁻¹, while the value of k₂′ (the composite of k₂K₆) is 2.36±0.05 mol⁻¹ dm³ s⁻¹ at 25.0 °C. In both cases there was reduction of the cobalt(III) centre to cobalt(II), but there was no reduction of the molybdenum(VI) centre. k₂₂, the self-exchange rate constant, for aqueous sulfite (as SO₃ ²⁻) was calculated to be 5.37×10⁻¹² mol⁻¹ dm³ s⁻¹, while for Fe(CN)₆ ⁴⁻, it was calculated to be 1.10×10⁹ mol⁻¹ dm³ s⁻¹ from the Marcus equations.