Reactivity of metal-oxo clusters towards biomolecules: from discrete polyoxometalates to metal-organic frameworks.

Metal-oxo clusters hold great potential in several fields such as catalysis, materials science, energy storage, medicine, and biotechnology. These nanoclusters of transition metals with oxygen-based ligands have also shown promising reactivity towards several classes of biomolecules, including proteins, nucleic acids, nucleotides, sugars, and lipids. This reactivity can be leveraged to address some of the most pressing challenges we face today, from fighting various diseases, such as cancer and viral infections, to the development of sustainable and environmentally friendly energy sources. For instance, metal-oxo clusters and related materials have been shown to be effective catalysts for biomass conversion into renewable fuels and platform chemicals. Furthermore, their reactivity towards biomolecules has also attracted interest in the development of inorganic drugs and bioanalytical tools. Additionally, the structural versatility of metal-oxo clusters allows for the efficiency and selectivity of the biomolecular reactions they promote to be readily tuned, thereby providing a pathway towards reaction optimization. The properties of the catalyst can also be improved through incorporation into solid supports or by linking metal-oxo clusters together to form Metal-Organic Frameworks (MOFs), which have been demonstrated to be powerful heterogeneous catalysts. Therefore, this review aims to provide a comprehensive and critical analysis of the state of the art on biomolecular transformations promoted by metal-oxo clusters and their applications, with a particular focus on structure-activity relationships.

Molecular Insights into Sequence-Specific Protein Hydrolysis by a Soluble Zirconium-Oxo Cluster Catalyst.

The development of catalysts for controlled fragmentation of proteins is a critical undertaking in modern proteomics and biotechnology. {Zr6O8}-based metal-organic frameworks (MOFs) have emerged as promising candidates for catalysis of peptide bond hydrolysis due to their high reactivity, stability, and recyclability. However, emerging evidence suggests that protein hydrolysis mainly occurs on the MOF surface, thereby questioning the need for their highly porous 3D nature. In this work, we show that the discrete and water-soluble [Zr6O4(OH)4(CH3CO2)8(H2O)2Cl3]+ (Zr6) metal-oxo cluster (MOC), which is based on the same hexamer motif found in various {Zr6O8}-based MOFs, shows excellent activity toward selective hydrolysis of equine skeletal muscle myoglobin. Compared to related Zr-MOFs, Zr6 exhibits superior reactivity, with near-complete protein hydrolysis after 24 h of incubation at 60 °C, producing seven selective fragments with a molecular weight in the range of 3-15 kDa, which are of ideal size for middle-down proteomics. The high solubility and molecular nature of Zr6 allow detailed solution-based mechanistic/interaction studies, which revealed that cluster-induced protein unfolding is a key step that facilitates hydrolysis. A combination of multinuclear nuclear magnetic resonance spectroscopy and pair distribution function analysis provided insight into the speciation of Zr6 and the ligand exchange processes occurring on the surface of the cluster, which results in the dimerization of two Zr6 clusters via bridging oxygen atoms. Considering the relevance of discrete Zr-oxo clusters as building blocks of MOFs, the molecular-level understanding reported in this work contributes to the further development of novel catalysts based on Zr-MOFs.

Green, Safe, and Reliable Synthesis of Bimetallic MOF-808 Nanozymes With Enhanced Aqueous Stability and Reactivity for Biological Applications.

Bimetallic metal-organic frameworks (MOFs) are promising nanomaterials whose reactivity towards biomolecules remains challenging due to issues related to synthesis, stability, control over metal oxidation state, phase purity, and atomic level characterization. Here, these shortcomings are rationally addressed through development of a synthesis of mixed metal Zr/Ce-MOFs in aqueous environment, overcoming significant hurdles in the development of MOF nanozymes, sufficiently stable on biologically relevant conditions. Specifically, a green and safe synthesis of Zr/Ce-MOF-808 is reported in water/acetic acid mixture which affords remarkably water-stable materials with reliable nanozymatic reactivity, including MOFs with a high Ce content previously reported to be unstable in water. The new materials outperform analogous bimetallic MOF nanozymes, showcasing that rational synthesis modifications could impart outstanding improvements. Further, atomic-level characterization by X-ray Absorption Fine Structure (XAFS) and X-ray Diffraction (XRD) confirmed superior nanozymes arise from differences in the synthetic method, which results in aqueous stable materials, and Ce incorporation, which perturbs the ligand exchange dynamics of the material, and could ultimately be used to fine tune the intrinsic MOF reactivity. Similar rational strategies which leverage metals in a synergistic manner should enable other water-stable bimetallic MOF nanozymes able to surpass existing ones, laying the path for varied biotechnological applications.

Supramolecular Self-Assembly of Proteins Promoted by Hybrid Polyoxometalates.

Controlling the formation of supramolecular protein assemblies and endowing them with new properties that can lead to novel functional materials is an important but challenging task. In this work, a new hybrid polyoxometalate is designed to induce controlled intermolecular bridging between biotin-binding proteins. Such bridging interactions lead to the formation of supramolecular protein assemblies incorporating metal-oxo clusters that go from several nanometers in diameter up to the micron range. Insights into the self-assembly process and the nature of the resulting biohybrid materials are obtained by a combination of Small Angle X-ray Scattering (SAXS), Transmission Electron Microscopy (TEM), and Dynamic Light Scattering (DLS), along with fluorescence, UV-vis, and Circular Dichroism (CD) spectroscopy. The formation of hybrid supramolecular assemblies is determined to be driven by biotin binding to the protein and electrostatic interactions between the anionic metal-oxo cluster and the protein, both of which also influence the stability of the resulting assemblies. As a result, the rate of formation, size, and stability of the supramolecular assemblies can be tuned by controlling the electrostatic interactions between the cluster and the protein (e.g., through varying the ionic strength of the solution), thereby paving the way toward biomaterials with tunable assembly and disassembly properties.

Monolacunary Wells-Dawson Polyoxometalate as a Novel Contrast Agent for Computed Tomography: A Comprehensive Study on In Vivo Toxicity and Biodistribution.

Polyoxotungstate nanoclusters have recently emerged as promising contrast agents for computed tomography (CT). In order to evaluate their clinical potential, in this study, we evaluated the in vitro CT imaging properties, potential toxic effects in vivo, and tissue distribution of monolacunary Wells-Dawson polyoxometalate, α2-K10P2W17O61.20H2O (mono-WD POM). Mono-WD POM showed superior X-ray attenuation compared to other tungsten-containing nanoclusters (its parent WD-POM and Keggin POM) and the standard iodine-based contrast agent (iohexol). The calculated X-ray attenuation linear slope for mono-WD POM was significantly higher compared to parent WD-POM, Keggin POM, and iohexol (5.97 ± 0.14 vs. 4.84 ± 0.05, 4.55 ± 0.16, and 4.30 ± 0.09, respectively). Acute oral (maximum-administered dose (MAD) = 960 mg/kg) and intravenous administration (1/10, 1/5, and 1/3 MAD) of mono-WD POM did not induce unexpected changes in rats' general habits or mortality. Results of blood gas analysis, CO-oximetry status, and the levels of electrolytes, glucose, lactate, creatinine, and BUN demonstrated a dose-dependent tendency 14 days after intravenous administration of mono-WD POM. The most significant differences compared to the control were observed for 1/3 MAD, being approximately seventy times higher than the typically used dose (0.015 mmol W/kg) of tungsten-based contrast agents. The highest tungsten deposition was found in the kidney (1/3 MAD-0.67 ± 0.12; 1/5 MAD-0.59 ± 0.07; 1/10 MAD-0.54 ± 0.05), which corresponded to detected morphological irregularities, electrolyte imbalance, and increased BUN levels.

Cavity-Directed Synthesis of Labile Polyoxometalates for Catalysis in Confined Spaces.

The artificial microenvironments inside coordination cages have gained significant attention for performing enzyme-like catalytic reactions by facilitating the formation of labile and complex molecules through a "ship-in-a-bottle" approach. Despite many fascinating examples, this approach remains scarcely explored in the context of synthesizing metallic clusters such as polyoxometalates (POMs). The development of innovative approaches to control and influence the speciation of POMs in aqueous solutions would greatly advance their applicability and could ultimately lead to the formation of elusive clusters that cannot be synthesized by using traditional methods. In this study, we employ host-guest stabilization within a coordination cage to enable a novel cavity-directed synthesis of labile POMs in aqueous solutions under mild conditions. The elusive Lindqvist [M6O19]2- (M=Mo or W) POMs were successfully synthesized at room temperature via the condensation of molybdate or tungstate building blocks within the confined cavity of a robust and water-soluble Pt6L4(NO3)12 coordination cage. Importantly, the encapsulation of these POMs enhances their stability in water, rendering them efficient catalysts for environmentally friendly and selective sulfoxidation reactions using H2O2 as a green oxidant in a pure aqueous medium. The approach developed in this paper offers a means to synthesize and stabilize the otherwise unstable metal-oxo clusters in water, which can broaden the scope of their applications.

Fine-tuning non-covalent interactions between hybrid metal-oxo clusters and proteins.

Interactions between the protein Hen Egg White Lysozyme (HEWL) and three different hybrid Anderson-Evans polyoxometalate clusters - AE-NH2 (δ-[MnMo6O18{(OCH2)3CNH2}2]3-), AE-CH3 (δ-[MnMo6O18{(OCH2)3CCH3}2]3-) and AE-Biot (δ-[MnMo6O18{(OCH2)3CNHCOC9H15N2OS}2]3-) - were studied via tryptophan fluorescence spectroscopy and single crystal X-ray diffraction. Quenching of tryptophan fluorescence was observed in the presence of all three hybrid polyoxometalate clusters (HPOMs), but the extent of quenching and the binding affinity were greatly dependent on the nature of the organic groups attached to the cluster. Control experiments further revealed the synergistic effect of the anionic polyoxometalate core and organic ligands towards enhanced protein interactions. Furthermore, the protein was co-crystallised with each of the three HPOMs, resulting in four different crystal structures, thus allowing for the binding modes of HPOM-protein interactions to be investigated with near-atomic precision. All crystal structures displayed a unique mode of binding of the HPOMs to the protein, with both functionalisation and the pH of the crystallisation conditions influencing the interactions. From the crystal structures, it was determined that HPOM-protein non-covalent complexes formed through a combination of electrostatic attraction between the polyoxometalate cluster and positively charged surface regions of HEWL, and direct and water-mediated hydrogen bonds with both the metal-oxo inorganic core and the functional groups of the ligand, where possible. Hence, functionalisation of metal-oxo clusters shows great potential in tuning their interactions with proteins, which is of interest for several biomedical applications.

Exploiting Interactions between Polyoxometalates and Proteins for Applications in (Bio)chemistry and Medicine.

The specific interactions of anionic metal-oxo clusters, known as polyoxometalates (POMs), with proteins can be leveraged for a wide range of analytical and biomedical applications. For example, POMs have been developed as selective catalysts that can induce protein modifications and have also been shown to facilitate protein crystallization, both of which are instrumental in the structural characterization of proteins. POMs can also be used for selective protein separation and enzyme inhibition, which makes them promising therapeutic agents. Hence, understanding POM-protein interactions is essential for the development of POM-based materials and their implementation in several fields. In this Review we summarize in detail the key insights that have been gained so far on POM-protein interactions. Emphasis is also given to hybrid POMs functionalized with organic ligands to prompt further research in this direction owing to the promising recent results on tuning POM-protein interactions through POM functionalization.

The Dawn of Metal-Oxo Clusters as Artificial Proteases: From Discovery to the Present and Beyond.

The selective cleavage of peptide bonds in proteins is of paramount importance in many areas of the biological and medical sciences, playing a key role in protein structure/function/folding analysis, protein engineering, and targeted proteolytic drug design. Current applications that depend on selective protein hydrolysis largely rely on costly proteases such as trypsin, which are sensitive to the pH, ionic strength, and temperature conditions. Moreover, >95% of peptides deposited in databases are generated from trypsin digests, restricting the information within the analyzed proteomes. On the other hand, harsh and toxic chemical reagents such as BrCN are very active but cause permanent modifications of certain amino acid residues. Consequently, transition-metal complexes have emerged as smooth and selective artificial proteases owing to their ability to provide larger fragments and complementary structural information. In the past decade, our group has discovered the unique protease activity of diverse metal-oxo clusters (MOC) and pioneered a distinctive approach to the development of selective artificial proteases. In contrast to classical coordination complexes which often depend on amino acid side chains to control the regioselectivity, the selectivity profile of MOCs is determined by a complex combination of structural factors, such as the protein surface charge, metal coordination to specific side chains, and hydrogen bonding between the protein surface and the MOC scaffold.In this Account, we present a critical overview of our detailed kinetic, spectroscopic, and crystallographic studies in MOC-assisted peptide bond hydrolysis, from its origins to the current rational and detailed mechanistic understanding. To this end, reactivity trends related to the structure and properties of MOCs based on the hydrolysis of small model peptides and key structural aspects governing the selectivity of protein hydrolysis are presented. Finally, our endeavors in seeking the next generation of heterogeneous MOC-based proteases are briefly discussed by embedding MOCs in metal-organic frameworks or using them as discrete nanoclusters in the development of artificial protease-like materials (i.e., nanozymes). The deep and comprehensive understanding sought experimentally and theoretically over the years in aqueous systems with intrinsic polar and charged substrates provides a unique view of the reactivity between inorganic moieties and biomolecules, thereby broadly impacting several different fields (e.g., catalysis in biochemistry, inorganic chemistry, and organic chemistry).

Expanding the Scope of Polyoxometalates as Artificial Proteases towards Hydrolysis of Insoluble Proteins.

Despite the enormous importance of insoluble proteins in biological processes, their structural investigation remains a challenging task. The development of artificial enzyme-like catalysts would greatly facilitate the elucidation of their structure since currently used enzymes in proteomics largely lose activity in the presence of surfactants, which are necessary to solubilize insoluble proteins. In this study, the hydrolysis of a fully insoluble protein by polyoxometalate complexes as artificial proteases in surfactant solutions is reported for the first time. The hydrolysis of zein as a model protein was investigated in the presence of Zr(IV) and Hf(IV) substituted Keggin-type polyoxometalates (POMs), (Et2 NH2 )10 [M(α-PW11 O39 )2 ] (M = Zr or Hf), and different concentrations of the anionic surfactant sodium dodecyl sulfate (SDS). Selective hydrolysis of the protein upon incubation with the catalyst was observed, and the results indicate that the hydrolytic selectivity and activity of the POM catalysts strongly depends on the concentration of surfactant. The molecular interactions between the POM catalyst and zein in the presence of SDS were explored using a combination of spectroscopic techniques which indicated competitive binding between POM and SDS towards the protein. Furthermore, the formation of micellar superstructures in ternary POM/surfactant/protein solutions has been confirmed by conductivity and Dynamic Light Scattering measurements.

High-resolution contrast-enhanced microCT reveals the true three-dimensional morphology of the murine placenta.

Genetic engineering of the mouse genome identified many genes that are essential for embryogenesis. Remarkably, the prevalence of concomitant placental defects in embryonic lethal mutants is highly underestimated and indicates the importance of detailed placental analysis when phenotyping new individual gene knockouts. Here we introduce high-resolution contrast-enhanced microfocus computed tomography (CE-CT) as a nondestructive, high-throughput technique to evaluate the 3D placental morphology. Using a contrast agent, zirconium-substituted Keggin polyoxometalate (Zr-POM), the soft tissue of the placenta (i.e., different layers and cell types and its vasculature) was imaged with a resolution of 3.5 µm voxel size. This approach allowed us to visualize and study early and late stages of placental development. Moreover, CE-CT provides a method to precisely quantify placental parameters (i.e., volumes, volume fraction, ratio of different placental layers, and volumes of specific cell populations) that are crucial for statistical comparison studies. The CE-CT assessment of the 3D morphology of the placentas was validated (i) by comparison with standard histological studies; (ii) by evaluating placentas from 2 different mouse strains, 129S6 and C57BL/6J mice; and (iii) by confirming the placental phenotype of mice lacking phosphoinositol 3-kinase (PI3K)-p110α. Finally, the Zr-POM-based CE-CT allowed for inspection of the vasculature structure in the entire placenta, as well as detecting placental defects in pathologies characterized by embryonic resorption and placental fusion. Taken together, Zr-POM-based CE-CT offers a quantitative 3D methodology to investigate placental development or pathologies.

Enhancing the Catalytic Activity of MOF-808 Towards Peptide Bond Hydrolysis through Synthetic Modulations.

The performance of MOFs in catalysis is largely derived from structural features, and much work has focused on introducing structural changes such as defects or ligand functionalisation to boost the reactivity of the MOF. However, the effects of different parameters chosen for the synthesis on the catalytic reactivity of the resulting MOF remains poorly understood. Here, we evaluate the role of metal precursor on the reactivity of Zr-based MOF-808 towards hydrolysis of the peptide bond in the glycylglycine model substrate. In addition, the effect of synthesis temperature and duration has been investigated. Surprisingly, the metal precursor was found to have a large influence on the reactivity of the MOF, surpassing the effect of particle size or number of defects. Additionally, we show that by careful selection of the Zr-salt precursor and temperature used in MOF syntheses, equally active MOF catalysts could be obtained after a 20 minute synthesis compared to 24 h synthesis.

Solution Dynamics of Hybrid Anderson-Evans Polyoxometalates.

Understanding the stability and speciation of metal-oxo clusters in solution is essential for many of their applications in different areas. In particular, hybrid organic-inorganic polyoxometalates (HPOMs) have been attracting increasing attention as they combine the complementary properties of organic ligands and metal-oxygen nanoclusters. Nevertheless, the speciation and solution behavior of HPOMs have been scarcely investigated. Hence, in this work, a series of HPOMs based on the archetypical Anderson-Evans structure, δ-[MnMo6O18{(OCH2)3C-R}2]3-, with different functional groups (R = -NH2, -CH3, -NHCOCH2Cl, -N═CH(2-C5H4N) {pyridine; -Pyr}, and -NHCOC9H15N2OS {biotin; -Biot}) and countercations (tetrabutylammonium {TBA}, Li, Na, and K) were synthesized, and their solution behavior was studied in detail. In aqueous solutions, decomposition of HPOMs into the free organic ligand, [MoO4]2-, and free Mn3+ was observed over time and was shown to be highly dependent on the pH, temperature, and nature of the ligand functional group but largely independent of ionic strength or the nature of the countercation. Furthermore, hydrolysis of the amide and imine bonds often present in postfunctionalized HPOMs was also observed. Hence, HPOMs were shown to exhibit highly dynamic behavior in solution, which needs to be carefully considered when designing HPOMs, particularly for biological applications.

Hybrid polyoxometalates as post-functionalization platforms: from fundamentals to emerging applications.

Polyoxometalates (POMs) represent an important group of metal-oxo nanoclusters, typically comprised of early transition metals in high oxidation states (mainly V, Mo and W). Many plenary POMs exhibit good pH, solvent, thermal and redox stability, which makes them attractive components for the design of covalently integrated hybrid organic-inorganic molecules, herein referred to as hybrid-POMs. Until now, thousands of organic hybrid-POMs have been reported; however, only a small fraction can be further functionalized using other organic molecules or metal cations. This emerging class of 'post-functionalizable' hybrid-POMs constitute a valuable modular platform that permits coupling of POM properties with different organic and metal cation functionalities, thereby expanding the key physicochemical properties that are relevant for application in (photo)catalysis, bioinorganic chemistry and materials science. The post-functionalizable hybrid-POM platforms offer an opportunity to covalently link multi-electron redox responsive POM cores with virtually any (bio)organic molecule or metal cation, generating a wide range of materials with tailored properties. Over the past few years, these materials have been showcased in the preparation of framework materials, functional surfaces, surfactants, homogeneous and heterogeneous catalysts and light harvesting materials, among others. This review article provides an overview on the state of the art in POM post-functionalization and highlights the key design and structural features that permit the discovery of new hybrid-POM platforms. In doing so, we aim to make the subject more comprehensible, both for chemists and for scientists with different materials science backgrounds interested in the applications of hybrid (POM) materials. The review article goes beyond the realms of polyoxometalate chemistry and encompasses emerging research domains such as reticular materials, surfactants, surface functionalization, light harvesting materials, non-linear optics, charge storing materials, and homogeneous acid-base catalysis among others.

Hydrolysis of Peptide Bonds in Protein Micelles Promoted by a Zirconium(IV)-Substituted Polyoxometalate as an Artificial Protease.

The development of artificial proteases is challenging, but important for many applications in modern proteomics and biotechnology. The hydrolysis of hydrophobic or unstructured proteins is particularly difficult due to their poor solubility, which often requires the presence of surfactants. Herein, it is shown that a zirconium(IV)-substituted Keggin polyoxometalate (POM), (Et2 NH2 )10 [Zr(α-PW11 O39 )2 ] (1), is able to selectively hydrolyze ß-casein, which is an intrinsically unstructured protein at pH 7.4 and 60 °C. Four surfactants (sodium dodecyl sulfate (SDS), N-dodecyl-N,N-dimethyl-3-ammonio-1-propanesulfonate (ZW3-12), 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS), and polyethylene glycol tert-octylphenyl ether (TX-100)), which differ in the nature of their polar groups, were investigated for their role in influencing the selectivity and efficiency of protein hydrolysis. Under experimental conditions, ß-casein forms micellar structures in which the hydrophilic part of the protein is water accessible and able to interact with 1. Identical fragmentation patterns of ß-casein in the presence of 1 were observed through SDS poly(acrylamide) gel electrophoresis both in the presence and absence of surfactants, but the rate of hydrolysis varied, depending on the nature of surfactant. Whereas TX-100 surfactant, which has a neutral polar head, caused only a slight decrease in the hydrolysis rate, stronger inhibition was observed in the presence surfactants with charges in their polar heads (CHAPS, ZW3-12, SDS). These results were consistent with those of tryptophan fluorescencequenching studies, which showed that the binding between ß-casein and 1 decreased with increasing repulsion between the POM and the polar heads of the surfactants. In all cases, the micellar structure of ß-casein was not significantly affected by the presence of POM or surfactants, as indicated by circular dichroism spectroscopy.

Selective Hydrolysis of Terminal Glycosidic Bond in α-1-Acid Glycoprotein Promoted by Keggin and Wells-Dawson Type Heteropolyacids.

The reactivity of a range of Keggin and Wells-Dawson type heteropolyacids (HPAs): H3 PW12 O40 H4 SiW12 O40 , H3 PMo12 O40 , K6 P2 W18 O62 , and NaH2 W12 O4 , towards the heavily glycosylated α-1-acid glycoprotein (AGP) is reported. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and high-performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD) show that after incubation of the protein with HPAs at 80 °C and pH 2.8 complete hydrolysis of terminal glycosidic bond has been achieved, resulting in the removal of sialic acids with no observed destruction of the protein core or the residual glycan chains. The 1 H NMR spectroscopy confirmed that the released sialic acids preserve intact structure upon their excision from the protein, which makes the reported method suitable for the analysis of sialic acid modifications which play an important role in numerous biological processes. The presence of other sugars was not detected by 1 H NMR and HPAEC-PAD, suggesting that HPAs hydrolyze only the terminal glycosidic bond in the glycoprotein, resulting in the selective release of sialic acid from AGP. The kinetic results have shown that under equal temperature and pH conditions, the hydrolysis of the terminal glucosidic bond occurred faster in the presence of HPAs compared to conventional mineral acids. The observed rate constants were in the range 6,7×10-2 -11,9×10-2  min-1 and the complete and selective excision of sialic acids could be achieved within 60 min of incubation. The Trp fluorescence and CD spectroscopy show that non-covalent interaction between HPA and protein takes place in solution which could lead to stabilization of the sialosyl cation that is formed during the glycosidic bond hydrolysis by anionic HPA cluster.

A Bis-organosilyl-Functionalized Wells-Dawson Polyoxometalate as a Platform for Facile Amine Postfunctionalization.

The development of modular platforms that can undergo postfunctionalization reactions permits coupling of inorganic clusters with different organic functionalities, thereby expanding the range of key physicochemical properties that are relevant for applications in different areas of science. In this work, a novel hybrid Wells-Dawson polyoxometalate (POM) platform was developed and successfully used for postfunctionalization via a nucleophilic substitution reaction. Two new halogen-functionalized bis-organosilyl Wells-Dawson POMs TBA6[α2-P2W17O61{O(SiC3H6-X)2}] (X = Cl or I) were synthesized, and their coupling with amine substrates was explored in a one-step postfunctionalization reaction. The iodide form of the POM has proven to be much more reactive, and its reaction with a range of primary and secondary amines resulted in a series of new bis-substituted Wells-Dawson POMs with the general formula TBA6[α2-P2W17O61{O(SiC3H6-NR1R2)2}]. Coupling of 18 amines with R1 and R2 groups, which exhibited a wide variety in terms of both chemical nature and bulkiness, was achieved under mild conditions via a catalyst-free approach. Using Na2CO3 as a base in acetonitrile solutions at 55 °C resulted in hybrid products that were obtained in high purity and good yields, after a simple isolation and purification procedure.

Redox Activity of Ce(IV)-Substituted Polyoxometalates toward Amino Acids and Peptides.

Redox reactions between polyoxometalates (POMs) and biologically relevant molecules have been virtually unexplored but are important, considering the growing interest in the biological applications of POMs. In this work we give a detailed account on the redox behavior of CeIV-substituted polyoxometalates (CeIV-POMs) toward a range of amino acids and peptides. CeIV-POMs have been shown to act as artificial proteases that promote the selective hydrolysis of peptide bonds. In presence of a protein, a concomitant reduction of CeIV to CeIII ion is frequently observed, leading us to examine the origins of this redox reaction by first using amino acid building blocks as simple models. Among all of the examined amino acids, cysteine (Cys) showed the highest activity in reducing CeIV-POMs to CeIII-POMs, followed by the aromatic amino acids tryptophan (Trp), tyrosine (Tyr), histidine (His), and phenylalanine (Phe). While the redox reaction with Cys afforded the well-defined product cystine, no oxidation products were detected for the Trp, His, Tyr, and Phe amino acids after their reaction with CeIV-POMs, suggesting a radical pathway in which the solvent likely regenerates the amino acid. In general, the rate of redox reactions increased upon increasing the pD, temperature, and ionic strength of the reaction. Moreover, the redox reaction is highly sensitive to the type of polyoxometalate scaffold, as complexation of CeIV to a Keggin (K) or Wells-Dawson (WD) polyoxotungstate anion resulted in a large difference in the rate of redox reaction for both Cys and aromatic amino acids. The reduction of CeIVK was at least 1 order of magnitude faster in comparison to CeIVWD, in accordance with the higher redox potential of CeIVK in comparison to CeIVWD. The reaction of CeIVPOMs with a range of peptides containing redox-active amino acids revealed that the redox reaction is influenced by their coordination mode with CeIV ion, but in all examined peptides the redox reaction is favored in comparison to the hydrolytic cleavage of the peptide bond.

Mechanism of the highly effective peptide bond hydrolysis by MOF-808 catalyst under biologically relevant conditions.

Efficient and selective hydrolysis of inert peptide bonds is of paramount importance. MOF-808, a metal-organic framework based on Zr6 nodes, can hydrolyze peptide bonds efficiently under biologically relevant conditions. However, the details of the catalyst structure and of the underlying catalytic reaction mechanism are challenging to establish. By means of DFT calculations we first investigate the speciation of the Zr6 nodes and identify the nature of ligands that bind to the Zr6O8H4-x core in aqueous conditions. The core is predicted to strongly prefer a Zr6O8H4 protonation state and to be predominantly decorated by bridging formate ligands, giving Zr6(µ3-O)4(µ3-OH)4(BTC)2(HCOO)6 and Zr6(µ3-O)4(µ3-OH)4(BTC)2(HCOO)5(OH)(H2O) as the most favorable structures at physiological pH. The GlyGly peptide can bind MOF in several different ways, with the preferred structure involving coordination through the terminal carboxylate analogously to the binding mode of formate ligand. The pre-reactive binding mode in which the amide carbonyl oxygen coordinates the metal core lies 7 kcal higher in free energy. The preferred reaction pathway is predicted to have two close-lying transition states, either of which could be the rate-determining step: nucleophilic attack on the amide carbon atom and C-N bond breaking, with calculated relative free energies of 31 and 32 kcal mol-1, respectively. Replacement of formate by water and hydroxide at the Zr6 node is predicted to be possible, but does not appear to play a role in the hydrolysis mechanism.

Selective Hydrolysis of Transferrin Promoted by Zr-Substituted Polyoxometalates.

The hydrolysis of the iron-binding blood plasma glycoprotein transferrin (Tf) has been examined at pH = 7.4 in the presence of a series of Zr-substituted polyoxometalates (Zr-POMs) including Keggin (Et2NH2)10[Zr(PW11O39)2]∙7H2O (Zr-K 1:2), (Et2NH2)8[{α-PW11O39Zr-(µ-OH) (H2O)}2]∙7H2O (Zr-K 2:2), Wells-Dawson K15H[Zr(α2-P2W17O61)2]·25H2O (Zr-WD 1:2), Na14[Zr4(α-P2W16O59)2(µ3-O)2(µ-OH)2(H2O)4]·57H2O (Zr-WD 4:2) and Lindqvist (Me4N)2[ZrW5O18(H2O)3] (Zr-L 1:1), (nBu4N)6[(ZrW5O18(µ-OH))2]∙2H2O (Zr-L 2:2)) type POMs. Incubation of transferrin with Zr-POMs resulted in formation of 13 polypeptide fragments that were observed on sodium dodecyl sulfate poly(acrylamide) gel electrophoresis (SDS-PAGE), but the hydrolysis efficiency varied depending on the nature of Zr-POMs. Molecular interactions between Zr-POMs and transferrin were investigated by using a range of complementary techniques such as tryptophan fluorescence, circular dichroism (CD), 31P-NMR spectroscopy, in order to gain better understanding of different efficiency of investigated Zr-POMs. A tryptophan fluorescence quenching study revealed that the most reactive Zr-WD species show the strongest interaction toward transferrin. The CD results demonstrated that interaction of Zr-POMs and transferrin in buffer solution result in significant secondary structure changes. The speciation of Zr-POMs has been followed by 31P-NMR spectroscopy in the presence and absence of transferrin, providing insight into stability of the catalysts under reaction condition.