JBIC Journal of Biological Inorganic Chemistry (v.22, #4)

Reactions of aquacobalamin and cob(II)alamin with chlorite and chlorine dioxide by Ilia A. Dereven’kov; Nikita I. Shpagilev; László Valkai; Denis S. Salnikov; Attila K. Horváth; Sergei V. Makarov (453-459).
Reactions of aquacobalamin (H2O–Cbl(III)) and its one-electron reduced form (cob(II)alamin, Cbl(II)) with chlorite (ClO2 ) and chlorine dioxide (ClO 2 ) were studied by conventional and stopped-flow UV–Vis spectroscopies and matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS). ClO2 does not react with H2O–Cbl(III), but oxidizes Cbl(II) to H2O–Cbl(III) as a major product and corrin-modified species as minor products. The proposed mechanism of chlorite reduction involves formation of OCl that modifies the corrin ring during the course of reaction with Cbl(II). H2O–Cbl(III) undergoes relatively slow destruction by ClO 2 via transient formation of oxygenated species, whereas reaction between Cbl(II) and ClO 2 proceeds extremely rapidly and leads to the oxidation of the Co(II)-center.
Keywords: Vitamin B12 ; Chlorite; Chlorine dioxide; Redox reactions; Kinetics

Synthesis of Ni(II) complexes bearing indole-based thiosemicarbazone ligands for interaction with biomolecules and some biological applications by Jebiti Haribabu; Kumaramangalam Jeyalakshmi; Yuvaraj Arun; Nattamai S. P. Bhuvanesh; Paramasivan Thirumalai Perumal; Ramasamy Karvembu (461-480).
A series of new Ni(II) complexes containing indole-based thiosemicarbazone ligands was synthesized and characterized by elemental analyses, and UV–visible, FT-IR, 1H & 13C NMR and mass spectroscopic techniques. The Ni(II) complexes (14) bear the general formula [Ni{C10H9N2NHCSNH(R)}2] where R = hydrogen (1), 4-methyl (2), 4-phenyl (3) and 4-cyclohexyl (4). Molecular structure of ligands (L3 and L4) and complexes (2, 3 and 4) was confirmed by single crystal X-ray crystallography. Four coordinated Ni(II) complexes showed square planar geometry. The interaction of the Ni(II) complexes with calf thymus DNA (CT-DNA) has been evaluated by absorption spectroscopic and ethidium bromide (EB) competitive binding studies, which revealed the intercalative interaction of the complexes with CT-DNA. Gel electrophoresis experiments showed the cleavage of DNA by the complexes without any external agent. Further, the interaction of the complexes with bovine serum albumin (BSA) was investigated using UV–visible, fluorescence and synchronous fluorescence spectroscopic methods, which showed that the complexes could bind strongly with BSA. Molecular docking was employed to understand the binding of the Ni(II) complexes with the molecular target B-DNA, human DNA topoisomerase I and BSA. All the Ni(II) complexes possess high antioxidant activity against 2-2-diphenyl-1-picrylhydrazyl (DPPH) radical and antihaemolytic activity. In addition, in vitro cytotoxicity of the Ni(II) complexes against lung cancer (A549), human breast cancer (MCF7) and mouse embryonic fibroblasts (L929) cell lines was investigated. Complex 4 has high cytotoxicity. The mode of cell death effected by complex 4 has been explored using Hoechst 33258 staining.Nickel(II) complexes of thiosemicarbazone ligands were synthesized and their DNA/protein binding, DNA cleavage and cytotoxicity abilities were studied.
Keywords: Indole; Thiosemicarbazones; Nickel(II) complexes; DNA/protein interactions; Cytotoxicity

EPR interpretation, magnetism and biological study of a Cu(II) dinuclear complex assisted by a schiff base precursor by Kuheli Das; Chiranjit Patra; Chandana Sen; Amitabha Datta; Chiara Massera; Eugenio Garribba; Mohamed Salah El Fallah; Belete B. Beyene; Chen-Hsiung Hung; Chittaranjan Sinha; Tulin Askun; Pinar Celikboyun; Daniel Escudero; Antonio Frontera (481-495).
A new Cu(II) dinuclear complex, Cu2L2 (1) was afforded employing the potentially pentatentate Schiff base precursor H2L, a refluxed product of o-vanillin and diethylenetriamine in methanol. Complex 1 was systematically characterized by FTIR, UV–Vis, emission and EPR spectrometry. The single crystal X-ray diffraction analysis of 1 reveals that the copper atom exhibits a distorted square planar geometry, comprising two pairs of phenolato-O and imine-N donors from two different H2L ligands. The temperature dependent magnetic interpretation agrees with the existence of weak antiferromagnetic interactions between the bridging dinuclear Cu(II) ions. A considerable body of experimental evidence has been accumulated to elucidate the magneto-structural relationship in this dinuclear Cu(II) complex by DFT computation. Both the ligand and complex 1 exhibit anti-mycobacterial activity and considerable efficacy on M. tuberculosis H37Ra (ATCC 25177) and M. tuberculosis H37Rv (ATCC 25618) strains. The practical applicability of the ligand and complex 1 has been examined in living cells (African Monkey Vero Cells). The MTT assay proves the non-toxicity of the probe up to 100 mg mL−1.A new homometallic dinuclear Cu(II) complex is afforded with a tetradentate Schiff base precursor. EPR interpretation and temperature dependent magnetic studies show that complex 1 has weak antiferromagnetic coupling and DFT computation is governed to explain the magneto-structural correlation.
Keywords: Cu(II) complex; Spectra; Magnetism; Anti-mycobacterial; Cytotoxicity

Non-thiolate ligation of nickel by nucleotide-free UreG of Klebsiella aerogenes by Vlad Martin-Diaconescu; Crisjoe A. Joseph; Jodi L. Boer; Scott B. Mulrooney; Robert P. Hausinger; Michael J. Maroney (497-503).
Nickel-dependent ureases are activated by a multiprotein complex that includes the GTPase UreG. Prior studies showed that nucleotide-free UreG from Klebsiella aerogenes is monomeric and binds one nickel or zinc ion with near-equivalent affinity using an undefined binding site, whereas nucleotide-free UreG from Helicobacter pylori selectively binds one zinc ion per dimer via a universally conserved Cys-Pro-His motif in each protomer. Iodoacetamide-treated K. aerogenes UreG was nearly unaffected in nickel binding compared to non-treated sample, suggesting the absence of thiolate ligands to the metal. X-ray absorption spectroscopy of nickel-bound UreG showed the metal possessed four-coordinate geometry with all O/N donor ligands including one imidazole, thus confirming the absence of thiolate ligation. The nickel site in Strep-tag II-modified protein possessed six-coordinate geometry, again with all O/N donor ligands, but now including two or three imidazoles. An identical site was noted for the Strep-tag II-modified H74A variant, substituted in the Cys-Pro-His motif, ruling out coordination by this His residue. These results are consistent with metal binding to both His6 and a His residue of the fusion peptide in Strep-tagged K. aerogenes UreG. We conclude that the nickel- and zinc-binding site in nucleotide-free K. aerogenes UreG is distinct from that of nucleotide-free H. pylori UreG and does not involve the Cys-Pro-His motif. Further, we show the Strep-tag II can perturb metal coordination of this protein.
Keywords: Urease; Metallocenter assembly; Nickel binding; X-ray absorption spectroscopy

Inhibition of copper-mediated aggregation of human γD-crystallin by Schiff bases by Priyanka Chauhan; Sai Brinda Muralidharan; Anand Babu Velappan; Dhrubajyoti Datta; Sanjay Pratihar; Joy Debnath; Kalyan Sundar Ghosh (505-517).
Protein aggregation, due to the imbalance in the concentration of Cu2+ and Zn2+ ions is found to be allied with various physiological disorders. Copper is known to promote the oxidative damage of β/γ-crystallins in aged eye lens and causes their aggregation leading to cataract. Therefore, synthesis of a small-molecule ‘chelator’ for Cu2+ with complementary antioxidant effect will find potential applications against aggregation of β/γ-crystallins. In this paper, we have reported the synthesis of different Schiff bases and studied their Cu2+ complexation ability (using UV–Vis, FT-IR and ESI-MS) and antioxidant activity. Further based on their copper complexation efficiency, Schiff bases were used to inhibit Cu2+-mediated aggregation of recombinant human γD-crystallin (HGD) and β/γ-crystallins (isolated from cataractous human eye lens). Among these synthesized molecules, compound 8 at a concentration of 100 μM had shown ~95% inhibition of copper (100 μM)-induced aggregation. Compound 8 also showed a positive cooperative effect at a concentration of 5–15 μM on the inhibitory activity of human αA-crystallin (HAA) during Cu2+-induced aggregation of HGD. It eventually inhibited the aggregation process by additional ~20%. However, ~50% inhibition of copper-mediated aggregation of β/γ-crystallins (isolated from cataractous human eye lens) was recorded by compound 8 (100 μM). Although the reductive aminated products of the imines showed better antioxidant activity due to their lower copper complexing ability, they were found to be non-effective against Cu2+-mediated aggregation of HGD.
Keywords: Schiff base; Copper complex; γ-Crystallin; α-Crystallin; Cu2+-induced aggregation inhibition

The X-ray structure of human apo-S100Z has been solved and compared with that of the zebrafish calcium-bound S100Z, which is the closest in sequence. Human apo-S100A12, which shows only 43% sequence identity to human S100Z, has been used as template model to solve the crystallographic phase problem. Although a significant buried surface area between the two physiological dimers is present in the asymmetric unit of human apo-S100Z, the protein does not form the superhelical arrangement in the crystal as observed for the zebrafish calcium-bound S100Z and human calcium-bound S100A4. These findings further demonstrate that calcium plays a fundamental role in triggering quaternary structure formation in several S100s. Solving the X-ray structure of human apo-S100Z by standard molecular replacement procedures turned out to be a challenge and required trying different models and different software tools among which only one was successful. The model that allowed structure solution was that with one of the lowest sequence identity with the target protein among the S100 family in the apo state. Based on the previously solved zebrafish holo-S100Z, a putative human holo-S100Z structure has been then calculated through homology modeling; the differences between the experimental human apo and calculated holo structure have been compared to those existing for other members of the family.
Keywords: S100Z; EF-hand; Oligomerization; Calcium; Molecular replacement

A method of expression for an oxygen-tolerant group III alcohol dehydrogenase from Pyrococcus horikoshii OT3 by Chikanobu Sugimoto; Kouta Takeda; Yumi Kariya; Hirotoshi Matsumura; Masafumi Yohda; Hiroyuki Ohno; Nobuhumi Nakamura (527-534).
NAD(P)-dependent group III alcohol dehydrogenases (ADHs), well known as iron-activated enzymes, generally lose their activities under aerobic conditions due to their oxygen-sensitivities. In this paper, we expressed an extremely thermostable group III ADH from the hyperthermophilic archaeon Pyrococcus horikoshii OT3 (PhADH) heterologously in Escherichia coli. When purified from a culture medium containing nickel, the recombinant PhADH (Ni-PhADH) contained 0.85 ± 0.01 g-atoms of nickel per subunit. Ni-PhADH retained high activity under aerobic conditions (9.80 U mg−1), while the enzyme expressed without adding nickel contained 0.46 ± 0.01 g-atoms of iron per subunit and showed little activity (0.27 U mg−1). In the presence of oxygen, the activity of the Fe2+-reconstituted PhADH prepared from the Ni-PhADH was gradually decreased, whereas the Ni2+-reconstituted PhADH maintained enzymatic activity. These results indicated that PhADH with bound nickel ion was stable in oxygen. The activity of the Ni2+-reconstituted PhADH prepared from the expression without adding nickel was significantly lower than that from the Ni-PhADH, suggesting that binding a nickel ion to PhADH in this expression system contributed to protecting against inactivation during the expression and purification processes. Unlike other thermophilic group III ADHs, Ni-PhADH showed high affinity for NAD(H) rather than NADP(H). Furthermore, it showed an unusually high k cat value toward aldehyde reduction. The activity of Ni-PhADH for butanal reduction was increased to 60.7 U mg−1 with increasing the temperature to 95 °C. These findings provide a new strategy to obtain oxygen-sensitive group III ADHs.
Keywords: Metalloenzyme; Alcohol dehydrogenase (ADH); Nickel; Extreme thermophile; Archaea

Urinary tract infections commonly occur in humans due to microbial pathogens invading the urinary tract, which can bring about a range of clinical symptoms and potentially fatal sequelae. The present study is aimed at addressing the development of a new antimicrobial agent against extended spectrum beta lactamase (ESBL) producing E. coli bacteria. We have synthesised some biologically potent (NNNN) donor macrocycles (L 1  = dibenzo[f,n]dipyrido[3,4-b:4′,3′-j][1,4,9,12]tetraazacyclohexadecine-6,11,18,23(5H,12H, 7H, 24H)-tetraone, and L 2  = 6,12,19,25-tetraoxo-4,6,11,12,16,18,23,24-octahydrotetrabenzo [b,g,k,p][1,5,10,14]tetra azacyclooctadecine-2,13-dicarboxylic acid) and their Ti and Zr metal complexes in alcoholic media using microwave protocol. Macrocyclic ligands were synthesised by incorporating of 3,5-diaminobenzoic acid, phthalic acid and 3,4-diaminopyridine in 1:1:1 molar ratio. The macrocyclic ligands and their metal complexes have been characterised by elemental analysis, conductance measurement, magnetic measurement and their structure configurations have been determined by various spectroscopic (FTIR, 1H/13C NMR, UV–Vis, LC–MS mass, XRD and TGA) techniques. [ZrL2Cl2]Cl2 metal complex shows excellent antibacterial activity against ESBLs. A zone of inhibition and minimum inhibitory concentration was determined by McFarland and the dilution method, respectively. The spectral studies confirm the binding sites of the nitrogen atom of the macrocycles. An octahedral geometry has been assigned to the metal complexes based on the findings.
Keywords: Macrocyclic ligand; Metal complexes; Spectral studies; ESBLs

Reactive sites and course of reduction in the Rieske protein by Si Ying Li; Paul H. Oyala; R. David Britt; Susan T. Weintraub; Laura M. Hunsicker-Wang (545-557).
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.
Keywords: Iron–sulfur cluster; Rieske proteins; DEPC; Circular dichroism; Mass spectrometry

Thiol oxidase ability of copper ion is specifically retained upon chelation by aldose reductase by Francesco Balestri; Roberta Moschini; Mario Cappiello; Umberto Mura; Antonella Del-Corso (559-565).
Bovine lens aldose reductase is susceptible to a copper-mediated oxidation, leading to the generation of a disulfide bridge with the concomitant incorporation of two equivalents of the metal and inactivation of the enzyme. The metal complexed by the protein remains redox active, being able to catalyse the oxidation of different physiological thiol compounds. The thiol oxidase activity displayed by the enzymatic form carrying one equivalent of copper ion (Cu1-AR) has been characterized. The efficacy of Cu1-AR in catalysing thiol oxidation is essentially comparable to the free copper in terms of both thiol concentration and pH effect. On the contrary, the two catalysts are differently affected by temperature. The specificity of the AR-bound copper towards thiols is highlighted with Cu1-AR being completely ineffective in promoting the oxidation of both low-density lipoprotein and ascorbic acid.
Keywords: Aldose reductase; Copper; Oxidative stress; Thiol oxidase

The exocyclic amino group of adenine in PtII and PdII complexes: a critical comparison of the X-ray crystallographic structural data and gas phase calculations by Radu Silaghi-Dumitrescu; Béla Mihály; Timea Mihály; Amr A. A. Attia; Pablo J. Sanz Miguel; Bernhard Lippert (567-579).
A detailed computational (DFT level of theory) study regarding the nature of the exocyclic amino group, N6H2, of the model nucleobase 9-methyladenine (9MeA) and its protonated (9MeAH+) and deprotonated forms (9MeA–H), free and metal-complexed, has been conducted. The metals are PtII and PdII, bonded to nitrogen-containing co-ligands (NH3, dien, bpy), with N1, N6, and N7 being the metal-binding sites, individually or in different combinations. The results obtained from gas phase calculations are critically compared with X-ray crystallography data, whenever possible. In the majority of cases, there is good qualitative agreement between calculated and experimentally determined C6–N6 bond lengths, but calculated values always show a trend to larger values, by 0.02–0.08 Å. Both methods indicate, with few exceptions, a high degree of double-bond character of C6–N6, consistent with an essentially sp2-hybridized N6 atom. The shortest values for C6–N6 distances in X-ray crystal structures are around 1.30 Å. Exceptions refer to cases in which DFT calculations suggest the existence of a hydrogen bond with N6H2 acting as a H bond acceptor, hence a situation with N6 having undergone a substantial hybridization shift toward sp3. Nevertheless, even in these cases the C6–N6 bond (1.392 Å) is still halfway between a typical C–N single bond (1.48 Å) and a typical C=N double bond (1.28 Å). This scenario is, however, not borne out by X-ray crystallographic results, and is attributed to the absence of counter anions and solvent molecules in the calculated structures.
Keywords: Adenine; Metal complexes; Nucleobase amino group; DFT calculations; Hybridization state

GHK and DAHK are biological peptides that bind both copper and zinc cations. Here we used infrared and Raman spectroscopies to study the coordination modes of both copper and zinc ions, at pH 6.8 and 8.9, correlating the data with the crystal structures that are only available for the copper-bound form. We found that Cu(II) binds to deprotonated backbone (amidate), the N-terminus and Nπ of the histidine side chain, in both GHK and DAHK, at pH 6.8 and 8.9. The data for the coordination of zinc at pH 6.8 points to two conformers including both nitrogens of a histidine residue. At pH 8.9, vibrational spectra of the ZnGHK complexes show that equilibria between monomers, oligomers exist, where deprotonated histidine residues as well as deprotonated amide nitrogen are involved in the coordination. A common feature is found: zinc cations coordinate to Nτ and/or Nπ of the His leading to the formation of GHK and DAHK multimers. In contrast, Cu(II) binds His via Nπ regardless of the peptide, in a pH-independent manner.
Keywords: Infrared spectroscopy; Raman spectroscopy; GHK; DAHK; Peptide copper coordination; Zinc coordination

EGFR-targeting peptide-coupled platinum(IV) complexes by Josef Mayr; Sonja Hager; Bettina Koblmüller; Matthias H. M. Klose; Katharina Holste; Britta Fischer; Karla Pelivan; Walter Berger; Petra Heffeter; Christian R. Kowol; Bernhard K. Keppler (591-603).
The high mortality rate of lung cancer patients and the frequent occurrence of side effects during cancer therapy demonstrate the need for more selective and targeted drugs. An important and well-established target for lung cancer treatment is the occasionally mutated epidermal growth factor receptor (EGFR). As platinum(II) drugs are still the most important therapeutics against lung cancer, we synthesized in this study the first platinum(IV) complexes coupled to the EGFR-targeting peptide LARLLT (and the shuffled RTALLL as reference). Notably, HPLC–MS measurements revealed two different peaks with the same molecular mass, which turned out to be a transcyclization reaction in the linker between maleimide and the coupled cysteine moiety. With regard to the EGFR specificity, subsequent biological investigations (3-day viability, 14-day clonogenic assays and platinum uptake) on four different cell lines with different verified EGFR expression levels were performed. Unexpectedly, the results showed neither an enhanced activity nor an EGFR expression-dependent uptake of our new compounds. Consequently, fluorophore-coupled peptides were synthesized to re-evaluate the targeting ability of LARLLT itself. However, also with these molecules, flow cytometry measurements showed no correlation of drug uptake with the EGFR expression levels. Taken together, we successfully synthesized the first platinum(IV) complexes coupled to an EGFR-targeting peptide; however, the biological investigations revealed that LARLLT is not an appropriate peptide for enhancing the specific uptake of small-molecule drugs into EGFR-overexpressing cancer cells.
Keywords: Platinum complexes; Anticancer drug; Peptides; EGFR

Effect of donor atom identity on metal-binding pharmacophore coordination by Benjamin L. Dick; Ashay Patel; J. Andrew McCammon; Seth M. Cohen (605-613).
The inhibition and binding of three metal-binding pharmacophores (MBPs), 2-hydroxycyclohepta-2,4,6-trien-1-one (tropolone), 2-mercaptopyridine-N-oxide (1,2-HOPTO), and 2-hydroxycyclohepta-2,4,6-triene-1-thione (thiotropolone) to human carbonic anhydrase II (hCAII) and a mutant protein hCAII L198G were investigated. These MBPs displayed bidentate coordination to the active site Zn(II) metal ion, but the MBPs respond to the mutation of L198G differently, as characterized by inhibition activity assays and X-ray crystallography. The L198G mutation increases the active site volume thereby decreasing the steric pressure exerted on MBPs upon binding, allowing changes in MBP coordination to be observed. When comparing the binding mode of tropolone to thiotropolone or 1,2-HOPTO (O,O versus O,S donor sets), structural modifications of the hCAII active site were shown to have a stronger effect on MBPs with an O,O versus O,S donor set. These findings were corroborated with density functional theory (DFT) calculations of model coordination complexes. These results suggest that the MBP binding geometry is a malleable interaction, particularly for certain ligands, and that the identity of the donor atoms influences the response of the ligand to changes in the protein active site environment. Understanding underlying interactions between a MBP and a metalloenzyme active site may aid in the design and development of potent metalloenzyme inhibitors.
Keywords: Computational chemistry; Density functional theory; Ligand binding; Metalloenzyme; X-ray crystallography

Computational evidence support the hypothesis of neuroglobin also acting as an electron transfer species by Licia Paltrinieri; Giulia Di Rocco; Gianantonio Battistuzzi; Marco Borsari; Marco Sola; Antonio Ranieri; Laura Zanetti-Polzi; Isabella Daidone; Carlo Augusto Bortolotti (615-623).
Neuroglobin (Ngb) is a recently identified hexa-coordinated globin, expressed in the nervous system of humans. Its physiological role is still debated: one hypothesis is that Ngb serves as an electron transfer (ET) species, possibly by reducing cytochrome c and preventing it to initiate the apoptotic cascade. Here, we use the perturbed matrix method (PMM), a mixed quantum mechanics/molecular dynamics approach, to investigate the redox thermodynamics of two neuroglobins, namely the human Ngb and GLB-6 from invertebrate Caenorhabditis elegans. In particular, we calculate the reduction potential of the two globins, resulting in an excellent agreement with the experimental values, and we predict the reorganization energies, λ, which have not been determined experimentally yet. The calculated λ values match well those reported for known ET proteins and thereby support a potential involvement in vivo of the two globins in ET processes.
Keywords: Hexa-coordinated globins; Reorganization energy; Molecular dynamics simulation; Cytochrome c ; Reduction potential; Thermodynamics

Remarkable reactivity of alkoxide/acetato-bridged binuclear copper(II) complex as artificial carboxylesterase by Bin Xu; Weidong Jiang; Xiaoqiang Liu; Fuan Liu; Zheng Xiang (625-635).
Bromo-containing binuclear Schiff base copper(II) complex, Cu2L(OAc), with an alkoxo/acetato-bridged moiety was employed as a model of carboxylesterases to promote the hydrolytic cleavage of p-nitrophenyl picolinate (PNPP). Furthermore, the reactivity of a mononuclear complex (CuHL) was evaluated for comparing it with that of binuclear one. The results reveal that the as-prepared binuclear Cu2L(OAc) efficiently accelerated the hydrolysis of PNPP, giving rise to excess four orders of magnitude rate enhancement in contrast to the un-catalyzed reaction. Cu2L(OAc) represented an enzyme-like bell-shaped pH-responsive kinetic behavior. Moreover, the binuclear one is more reactive than its mononuclear analogue (CuHL) by two orders of magnitude. The total efficiency of Cu2L(OAc) is about 61-fold than that of its mononuclear analogue, CuHL. In addition, a contrast experiment reveals that binuclear Cu2L(OAc) displayed good activity in the hydrolysis of PNPP as well another active ester, i.e., S-2-benzothiazolyl 2-amino-alpha-(methoxyimino)-4-thiazolethiolacetate (AE-active ester). Noteworthyly, it was found that mononuclear one inspired more obvious rate enhancement in the hydrolysis of AE-active ester relative to PNPP hydrolysis. The estimated pK a1 of bound water on the binuclear Cu2L(OAc) using second derivative method (SDM) is relatively smaller than that for CuHL by a gap of about 0.8 pK unit, which facilitates the hydrolysis of PNPP.Four orders of magnitude rate enhancement was observed for the catalytic hydrolysis of p-nitrophenyl picolinate (PNPP) by one μ-alkoxide/acetato-bridged binuclear copper(II) complex under physiological conditions. Substrate specificity of the resulting binuclear complexes was observed for the hydrolysis of PNPP and AE-active ester.
Keywords: Bimetallic cooperation; Artificial enzyme; Hydrolysis; Comparable kinetics