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.     

Guanidination of notexin alters its membrane-damaging activity in response to sphingomyelin and cholesterol

To elucidate the contribution of phospholipase A₂ (PLA₂) activity of notexin to its ability to perturb membranes, comparative studies on the interaction of notexin and guanidinated notexin (Gu-notexin) with egg yolk phosphatidylcholine (EYPC), EYPC/egg yolk sphingomyelin (EYSM) and EYPC/EYSM/cholesterol vesicles were conducted. EYSM notably reduced the membrane-damaging activity of notexin against EYPC vesicles, but had an insignificant influence on that of Gu-notexin. Unlike the effects noted with notexin, inactivation of PLA₂ activity by EDTA led to a reduction in the ability of Gu-notexin to induce EYPC/EYSM vesicle leakage and to increase Gu-notexin-induced membrane permeability of EYPC/EYSM/cholesterol vesicles. The geometrical arrangement of notexin and Gu-notexin in contact with either EYPC/EYSM vesicles or EYPC/EYSM/cholesterol vesicles differed. Moreover, global conformation of notexin and Gu-notexin differed in either Ca²⁺-bound or metal-free states. These results indicate that notexin and Gu-notexin could induce membrane permeability without the involvement of PLA₂ activity, and suggest that guanidination alters the membrane-bound mode of notexin on damaging phospholipid vesicles containing sphingomyelin and cholesterol.     

Inclusion complex of 2-chlorobenzophenone with cyclomaltoheptaose (β-cyclodextrin): temperature, solvent effects and molecular modeling

A thermodynamic study of the inclusion process between 2-chlorobenzophenone (2ClBP) and cyclomaltoheptaose (β-cyclodextrin, β-CD) was performed using UV–vis spectroscopy, reversed-phase liquid chromatography (RP-HPLC), and molecular modeling (PM6). Spectrophotometric measurements in aqueous solutions were performed at different temperatures. The stoichiometry of the complex is 1:1 and its apparent formation constant (K_c) is 3846 M⁻¹ at 30 °C. Temperature dependence of K_c values revealed that both enthalpy (ΔH° = −10.58 kJ/mol) and entropy changes (ΔS° = 33.76 J/K mol) are favorable for the inclusion process in an aqueous medium. Encapsulation was also investigated using RP-HPLC (C18 column) with different mobile-phase compositions, to which β-CD was added. The apparent formation constants in MeOH–H₂O (K_F) were dependent of the proportion of the mobile phase employed (50:50, 55:45, 60:40 and 65:35, v/v). The K_F values were 419 M⁻¹ (50% MeOH) and 166 M⁻¹ (65% MeOH) at 30 °C. The thermodynamic parameters of the complex in an aqueous MeOH medium indicated that this process is largely driven by enthalpy change (ΔH° = −27.25 kJ/mol and ΔS° = −45.12 J/K mol). The results of the study carried out with the PM6 semiempirical method showed that the energetically most favorable structure for the formation of the complex is the ‘head up’ orientation.     

Configuration of polyisoprenoids affects the permeability and thermotropic properties of phospholipid/polyisoprenoid model membranes

The influence of α-cis- and α-trans-polyprenols on the structure and properties of model membranes was analyzed. The interaction of Ficaprenol-12 (α-cis-Prenol-12, α-Z-Prenol-12) and Alloprenol-12 (α-trans-Prenol-12, α-E-Prenol-12) with model membranes was compared using high performance liquid chromatography (HPLC), differential scanning calorimetry (DSC) and fluorescent methods. l-α-Phosphatidylcholine from egg yolk (EYPC) and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) as the main lipid components of unilamellar (SUVs) and multilamellar (MLVs) vesicles were used. The two-step extraction procedure (n-pentane and hexane, respectively) allowed to separately analyze the fractions of polyprenol as non-incorporated (Prenol_NonInc) and incorporated (Prenol_Inc) into liposomes. Consequently, distribution coefficients, P′, describing the equilibrium of prenol content between phospholipid (EYPC) membrane and the aqueous phase gave different log P′ for α-cis- and α-trans-Prenol-12, indicating that the configuration of the α-terminal residue significantly alters the hydrophobicity of the polyisoprenoid molecule and consequently the affinity of polyprenols for EYPC membrane. In fluorescence experiments α-trans-Pren-12 increased up to 1.7-fold the permeability of EYPC bilayer for glucose while the effect of α-cis-Pren-12 was almost negligible. Considerable changes of thermotropic behavior of DPPC membranes in the presence of both prenol isomers were observed. α-trans-Pren-12 completely abolished the pretransition while in the case of α-cis-Pren-12 it was noticeably reduced. Furthermore, for both prenol isomers, the temperature of the main phase transition (T_m) was shifted by about 1 °C to lower values and the height of the peak was significantly reduced. The DSC analysis profiles also showed a new peak at 38.7 °C, which may suggest the concomitant presence of more that one phase within the membrane.Results of these experiments and the concomitant occurrence of alloprenols and ficaprenols in plant tissues suggest that cis/trans isomerization of the α-residue of polyisoprenoid molecule might comprise a putative mechanism responsible for modulation of the permeability of cellular membranes.     

Inactivation of α1-Proteinase Inhibitor as a Broad Screen for Detecting Proteolytic Activities in Unknown Samples

The need for a quick, simple screening method for the detection of general proteolytic activity prompted us to determine whether cleavage within the reactive site loop region (RSL) of α₁-proteinase inhibitor (α₁-PI), a well-characterized member of the serpin family known to be susceptible to proteolytic inactivation, can be utilized for this purpose. Inactivation of α₁-PI in the RSL region can be measured by loss of residual inhibitory capacity of α₁-PI against its target proteinase. While we originally utilized this assay to detect a new proteinase from culture supernatants ofPorphyromonas gingivalis, the feasibility of extending this assay to scan for proteolytic activity from other systems was also assessed. As an example, we found that the serine proteinase fromStaphylococcus aureus(SSP) had virtually the same catalytic efficiency in inactivating α₁-PI in our assay as it did in the hydrolysis of the synthetic substrate Z-Phe-Leu-Glu–pNA (k_cat/K_mvalue of 2 × 10⁴M⁻¹s⁻¹vs 2.6 × 10⁴M⁻¹s⁻¹, respectively). Additionally, in both assays activity could be readily detected in less than a 1 h incubation at SSP concentrations in the picomolar range. This assay is unique in that proteinases which hydrolyze peptide bonds within the RSL of α₁-PI can readily be detected as measured by loss of α₁-PI inhibitory activity.     

A study on the reaction of copper complex of dioxotetraamine with superoxide ion by spectrophotometry and pulse radiolysis

The reactions of a dioxotetraamine Cu(II) complex [Cu(H₋₂L)] (L is 6-(9-fluorenyl)-1,4,8,11-tetraazaandencane-5,7-dione)with O₂ ⁻ were investigated by electrochemistry, UV-Vis spectrophotometry and pulse radiolysis, respectively. In DMSO solution, [Cu^II(H₋₂L)] was oxidized into [Cu^III(H₋₂L)]⁺ by O₂ ⁻, a consecutive reaction was observed with [Cu^III(H₋₂L)(O₂ ²⁻)] ⁻ as intermediates (k₁=1.71×10³ M⁻¹ s⁻¹, k₂=1.2×10⁻² s⁻¹). The mechanism of O₂ ⁻ dismutation catalyzed by the complex involved alternate oxidation and reduction of Cu(II) by O₂ ⁻ and the k_cat is 6.07 × 10⁷ M⁻¹ s⁻¹ (pH 7.4).     

Diorganotin(IV) complexes of dipeptides containing the alpha-aminoisobutyryl residue (Aib): preparation, structural characterization, antibacterial and antiproliferative activities of [(n-Bu)2 Sn(H(-1)L)] (LH=H-Aib-L-Leu-OH, H-Aib-L-Ala-OH)

Two new organotin(IV) complexes with dianionic dipeptides containing the α-aminoisobutyryl residue (Aib) as ligands are described. The solid complexes [(n-Bu)₂Sn(H₋₁L_A)] · 2MeOH (1 · 2MeOH) (L_AH = H-Aib-L-Leu-OH) and [(n-Bu)₂Sn(H₋₁L_B)] · MeOH (2 · MeOH) (L_BH = H-Aib-L-Ala-OH) have been isolated and characterized by single-crystal X-ray crystallography and spectroscopic techniques (H₋₁L²⁻ is the dianionic form of the corresponding dipeptide). Complexes 1 · 2MeOH and 2 · MeOH are monomeric with similar molecular structures. The doubly deprotonated dipeptide behaves as a N(amino), N(peptide), O(carboxylate) ligand and binds to the Sn^IV atom. The five-coordinate metal ion has a distorted trigonal bipyramidal geometry. A different network of intermolecular hydrogen bonds in each compound results in very dissimilar supramolecular features. The IR, far-IR, Raman and ¹¹⁹Sn NMR data are discussed in terms of the nature of bonding and known structures. The antibacterial and antiproliferative activities as well as the effect of the new compounds on pDNA were examined. Complexes 1 and 2 are active against the gram-positive bacteria Bacillus subtilis and Bacillus cereus. The IC₅₀ values reveal that the two compounds express promising cytotoxic activity in vitro against a series of cell lines.     

Paraoxon, 4-nitrophenyl phosphate and acetate are substrates of α- but not of β-, γ- and ζ-carbonic anhydrases

Carbonic anhydrases (CAs, EC 4.2.1.1) belonging to α-, β-, γ- and ζ-classes and from various organisms, ranging from the bacteria, archaea to eukarya domains, were investigated for their esterase/phosphatase activity with 4-nitrophenyl acetate, 4-nitrophenyl phosphate and paraoxon as substrates. Only α-CAs showed esterase/phosphatase activity, whereas enzymes belonging to the β-, γ- and ζ-classes were completely devoid of such activity. Paraoxon, the metabolite of the organophosphorus insecticide parathione, was a much better substrate for several human/murine α-CA isoforms (CA I, II and XIII), with k_cat/K_M in the range of 2681.6–4474.9 M^?1 s^?1, compared to 4-nitrophenyl phosphate (k_cat/K_M of 14.9–1374.4 M^?1 s^?1).     

Structure and mechanism of a cysteine sulfinate desulfinase engineered on the aspartate aminotransferase scaffold

The joint substitution of three active-site residues in Escherichia colil-aspartate aminotransferase increases the ratio of l-cysteine sulfinate desulfinase to transaminase activity 10⁵-fold. This change in reaction specificity results from combining a tyrosine-shift double mutation (Y214Q/R280Y) with a non-conservative substitution of a substrate-binding residue (I33Q). Tyr214 hydrogen bonds with O3 of the cofactor and is close to Arg374 which binds the α-carboxylate group of the substrate; Arg280 interacts with the distal carboxylate group of the substrate; and Ile33 is part of the hydrophobic patch near the entrance to the active site, presumably participating in the domain closure essential for the transamination reaction. In the triple-mutant enzyme, k_cat′ for desulfination of l-cysteine sulfinate increased to 0.5 s^− 1 (from 0.05 s^− 1 in wild-type enzyme), whereas k_cat′ for transamination of the same substrate was reduced from 510 s^− 1 to 0.05 s^− 1. Similarly, k_cat′ for β-decarboxylation of l-aspartate increased from < 0.0001 s^− 1 to 0.07 s^− 1, whereas k_cat′ for transamination was reduced from 530 s^− 1 to 0.13 s^− 1. l-Aspartate aminotransferase had thus been converted into an l-cysteine sulfinate desulfinase that catalyzes transamination and l-aspartate β-decarboxylation as side reactions. The X-ray structures of the engineered l-cysteine sulfinate desulfinase in its pyridoxal-5′-phosphate and pyridoxamine-5′-phosphate form or liganded with a covalent coenzyme-substrate adduct identified the subtle structural changes that suffice for generating desulfinase activity and concomitantly abolishing transaminase activity toward dicarboxylic amino acids. Apparently, the triple mutation impairs the domain closure thus favoring reprotonation of alternative acceptor sites in coenzyme-substrate intermediates by bulk water.     

[H,N] NMR studies of the aquation of cis-diamine platinum(II) complexes

Two ¹⁵N-labelled cis-Pt(II) diamine complexes with dimethylamine (¹⁵N-dma) and isopropylamine (¹⁵N-ipa) ligands have been prepared and characterised. [¹H,¹⁵N] HSQC NMR spectroscopy is used to obtain the rate and equilibrium constants for the aquation of cis-[PtCl₂(¹⁵N-dma)₂] at 298 K in 0.1 M NaClO₄ and to determine the pK_a values of cis-[PtCl(H₂O)(¹⁵N-dma)₂]⁺ (6.37) and cis-[Pt(H₂O)₂(¹⁵N-dma)₂]²⁺ (pK_a1 = 5.17, pK_a2 = 6.47). The rate constants for the first and second aquation steps (k₁ = (2.12 ± 0.01) × 10⁻⁵ s⁻¹, k₂ = (8.7 ± 0.7) × 10⁻⁶ s⁻¹) and anation steps (k₋₁ = (6.7 ± 0.8) × 10⁻³ M⁻¹ s⁻¹, k₋₂ = 0.043 ± 0.004 M⁻¹ s⁻¹) are very similar to those reported for cisplatin under similar conditions, and a minor difference is that slow formation of the hydroxo-bridged dimer is observed. Aquation studies of cis-[PtCl₂(¹⁵N-ipa)₂] were precluded by the close proximity of the NH proton signal to the ¹H₂O resonance.     

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.     

Kinetic study of the quenching reaction of singlet oxygen by α-, β-, γ-, δ-tocotrienols,and palm oil and soybean extracts in solution

Measurements of the singlet oxygen (¹O₂) quenching rates (k_Q (S)) and the relative singlet oxygen absorption capacity (SOAC) values were performed for 11 antioxidants (AOs) (eight vitamin E homologues (α-, β-, γ-, and δ-tocopherols and -tocotrienols (-Tocs and -Toc-3s)), two vitamin E metabolites (α- and γ-carboxyethyl-6-hydroxychroman), and trolox) in ethanol/chloroform/D₂O (50:50:1, v/v/v) and ethanol solutions at 35?°C. Similar measurements were performed for five palm oil extracts 1–5 and one soybean extract 6, which included different concentrations of Tocs, Toc-3s, and carotenoids. Furthermore, the concentrations (wt%) of Tocs, Toc-3s, and carotenoids included in extracts 1–6 were determined. From the results, it has been clarified that the ¹O₂-quenching rates (k_Q (S)) (that is, the relative SOAC value) obtained for extracts 1–6 may be explained as the sum of the product {Σ k_Q^AO-i (S) [AO-i]/100} of the rate constant (k_Q^AO-i (S)) and the concentration ([AO-i]/100) of AO-i (Tocs, Toc-3s, and carotenoid) included.     

Trans-pyridine tetraammine complexes of RuII and RuIII with N7-coordninated purine nucleosides

 The synthesis, spectroscopic, and electrochemical properties of trans-[L(Pyr)(NH₃)₄Ru^II/III] (Pyr=py, 3-phpy, 4-phpy, 3-bnpy, or 4-bnpy; L=H₂O, Guo, dGuo, 1MeGuo, Gua, Ino, or G⁷-DNA) are reported. As expected, the Pyr ligand slows DNA binding by trans-[(H₂O)(Pyr)(NH₃)₄Ru^II]²⁺ relative to [(H₂O)(NH₃)₅Ru^II]²⁺ and favors reduction of Ru^III by about 150 mV. The pyridine ligand also promotes the disproportionation of Ru^III to afford the corresponding complexes of Ru^II and, presumably, Ru^IV. For L=Ino, disproportionation follows the rate law: d[Ru^II]/dt=k ₀[Ru^III]+k ₁[OH^–][Ru^III], k ₀=(2.7±0.7)×10^–4s^–1 and k ₁=70±1 M^–1s^–1. Received: 11 May 1998 / Accepted: 3 March 1999     

Aggregation of imino-phosphonate monoester complexes

The aggregates {[Zn(L¹)]H₂O}_∞ and {[Y(L²)]₄Na₃(H₂O)₂(MeOH)_1.2}(NO₃)₃·2H₂O·5.6MeOH have been assembled from complexes of imino-phosphonate monoester ligands [L¹]²⁻ {CH₂[CH₂NC(CH₃)PO₂(OMe)]₂}²⁻ and [L²]³⁻ {N[CH₂CH₂NC(CH₃)PO₂(OMe)]₃}³⁻, the topology of these materials differing from that of their imino-carboxylate analogues.     

Single-Molecule Kinetics Reveal Cation-Promoted DNA Duplex Formation Through Ordering of Single-Stranded Helices

In this work, the kinetics of short, fully complementary oligonucleotides are investigated at the single-molecule level. Constructs 6–9 bp in length exhibit single exponential kinetics over 2 orders of magnitude time for both forward (k_on, association) and reverse (k_off, dissociation) processes. Bimolecular rate constants for association are weakly sensitive to the number of basepairs in the duplex, with a 2.5-fold increase between 9 bp (k′_on = 2.1(1) × 10⁶ M⁻¹ s⁻¹) and 6 bp (k′_on = 5.0(1) × 10⁶ M⁻¹ s⁻¹) sequences. In sharp contrast, however, dissociation rate constants prove to be exponentially sensitive to sequence length, varying by nearly 600-fold over the same 9 bp (k_off = 0.024 s⁻¹) to 6 bp (k_off = 14 s⁻¹) range. The 8 bp sequence is explored in more detail, and the NaCl dependence of k_on and k_off is measured. Interestingly, k_onincreases by >40-fold (k_on = 0.10(1) s⁻¹ to 4.0(4) s⁻¹ between [NaCl] = 25 mM and 1 M), whereas in contrast, k_offdecreases by fourfold (0.72(3) s⁻¹ to 0.17(7) s⁻¹) over the same range of conditions. Thus, the equilibrium constant (K_eq) increases by ≈160, largely due to changes in the association rate, k_on. Finally, temperature-dependent measurements reveal that increased [NaCl] reduces the overall exothermicity (ΔΔH° > 0) of duplex formation, albeit by an amount smaller than the reduction in entropic penalty (−TΔΔS° < 0). This reduced entropic cost is attributed to a cation-facilitated preordering of the two single-stranded species, which lowers the association free-energy barrier and in turn accelerates the rate of duplex formation.     

Mechanism and biological implications of the NO release of cis-[Ru(bpy)2L(NO)](n+) complexes: a key role of physiological thiols

Nitric oxide (NO) has a critical role in several physiological and pathophysiological processes. In this paper, the reactions of the nitrosyl complexes of [Ru(bpy)₂L(NO)]ⁿ⁺ type, where L = SO₃²⁻ and imidazole and bpy = 2,2′-bipiridine, with cysteine and glutathione were studied. The reactions with cysteine and glutathione occurred through the formation of two sequential intermediates, previously described elsewhere, [Ru(bpy)₂L(NOSR)]ⁿ⁺ and [Ru(bpy)₂L(NOSR)₂] (SR = thiol) leading to the final products [Ru(bpy)₂L(H₂O)]ⁿ⁺ and free NO. The second order rate constant for the second step of this reaction was calculated for cysteine k₂(SR⁻) = (2.20 ± 0.12) × 10⁹ M^− 1 s^− 1 and k_2(RSH) = (154 ± 2) M^− 1 s^− 1 for L = SO₃²⁻ and k₂(SR⁻) = (1.30 ± 0.23) × 10⁹ M^− 1 s^− 1 and k₂(RSH) = (0.84 ± 0.02) M^− 1 s^− 1 for L = imidazole; while for glutathione they were k₂(SR⁻) = (6.70 ± 0.32) × 10⁸ M^− 1 s^− 1 and k₂(RSH) = 11.8 ± 0.3 M^− 1 s^− 1 for L = SO₃²⁻ and k₂(SR⁻) = (2.50 ± 0.36) × 10⁸ M^− 1 s^− 1 and k₂(RSH) = 0.32 ± 0.01 M^− 1 s^− 1 for L = imidazole. In all reactions it was possible to detect the release of NO from the complexes, which it is remarkably distinct from other ruthenium metallocompounds described elsewhere with just N₂O production. These results shine light on the possible key role of NO release mediated by physiological thiols in reaction with these metallonitrosyl ruthenium complexes.     

Biochemical characterization of recombinant β-carbonic anhydrase (PgiCAb) identified in the genome of the oral pathogenic bacterium Porphyromonas gingivalis

Abstract

Carbonic anhydrases (CAs, EC 4.2.1.1) belonging to the α-, β-, γ-, δ- and ζ-CAs are ubiquitous metalloenzymes present in prokaryotes and eukaryotes. CAs started to be investigated in detail only recently in pathogenic bacteria, in the search for antibiotics with a novel mechanism of action, since it has been demonstrated that in many such organisms they are essential for the life cycle of the organism. CA inhibition leads to growth impairment or growth defects in several pathogenic bacteria. The microbiota of the human oral mucosa consists of a myriad of bacterial species, Porphyromonas gingivalis being one of them and the major pathogen responsible for the development of chronic periodontitis. The genome of P. gingivalis encodes for a β- and a γ-CAs. Recently, our group purified the recombinant γ-CA (named PgiCA) which was shown to possess a significant catalytic activity for the reaction that converts CO₂ to bicarbonate and protons, with a k_cat of 4.1?×?10^5?s^?1 and a k_cat/K_m of 5.4?×?10^7?M^?1?×?s^?1. We have also investigated its inhibition profile with a range of inorganic anions such as thiocyanate, cyanide, azide, hydrogen sulfide, sulfamate and trithiocarbonate. Here, we describe the cloning, purification and kinetic parameters of the other class of CA identified in the genome of P. gingivalis, the β-CA, named PgiCAb. This enzyme has a good catalytic activity, with a k_cat of 2.8?×?10^5?s^?1 and a k_cat/K_m of 1.5?×?10^7?M^?1?×?s^?1. PgiCAb was also inhibited by the clinically used sulfonamide acetazolamide, with an inhibition constant of 214?nM. The role of CAs as possible virulence factors of P. gingivalis is poorly understood at the moment but their good catalytic activity and the fact that they might be inhibited by a large number of compounds, which may pave the way for finding inhibitors with antibacterial activity that may elucidate these phenomena and lead to novel antibiotics.     

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.