Mushroom Tyrosinase: Six Isoenzymes Catalyzing Distinct Reactions.

"Mushroom tyrosinase" from the common button mushroom is the most frequently used source of tyrosinase activity, both for basic and applied research. Here, the complete tyrosinase family from Agaricus bisporus var. bisporus (abPPO1-6) was cloned from mRNA and expressed heterologously using a single protocol. All six isoenzymes accept a wide range of phenolic and catecholic substrates, but display pronounced differences in their specificity and enzymatic reaction rate. AbPPO3 ignores γ-l-glutaminyl-4-hydroxybenzene (GHB), a natural phenol present in mM concentrations in A. bisporus, while AbPPO4 processes 100 µM GHB at 4-times the rate of the catechol l-DOPA. All six AbPPOs are biochemically distinct enzymes fit for different roles in the fungal life cycle, which challenges the traditional concept of isoenzymes as catalyzing the same physiological reaction and varying only in secondary properties. Transferring this approach to other enzymes and organisms will greatly stimulate both the study of the in vivo function(s) of enzymes and the application of these highly efficient catalysts.

Polyphenol exposure of mothers and infants assessed by LC-MS/MS based biomonitoring in breast milk.

Exposure to polyphenols is relevant throughout critical windows of infant development, including the breastfeeding phase. However, the quantitative assessment of polyphenols in human breast milk has received limited attention so far, though polyphenols may positively influence infant health. Therefore, a targeted LC-MS/MS assay was developed to investigate 86 analytes representing different polyphenol classes in human breast milk. The sample preparation consisted of liquid extraction, salting out, freeze-out, and a dilution step. Overall, nearly 70% of the chemically diverse polyphenols fulfilled all strict validation criteria for full quantitative assessment. The remaining analytes did not fulfill all criteria at every concentration level, but can still provide useful semi-quantitative insights into nutritional and biomedical research questions. The limits of detection for all analyzed polyphenols were in the range of 0.0041-87 ng*mL-1, with a median of 0.17 ng*mL-1. Moreover, the mean recovery was determined to be 82% and the mean signal suppression and enhancement effect was 117%. The developed assay was applied in a proof-of-principle study to investigate polyphenols in breast milk samples provided by twelve Nigerian mothers at three distinct time points post-delivery. In total, 50 polyphenol analytes were detected with almost half being phenolic acids. Phase II metabolites, including genistein-7-ß-D-glucuronide, genistein-7-sulfate, and daidzein-7-ß-D-glucuronide, were also detected in several samples. In conclusion, the developed method was demonstrated to be fit-for-purpose to simultaneously (semi-) quantify a wide variety of polyphenols in breast milk. It also demonstrated that various polyphenols including their biotransformation products were present in breast milk and therefore likely transferred to infants where they might impact microbiome development and infant health.

Exposomic Biomonitoring of Polyphenols by Non-Targeted Analysis and Suspect Screening.

Polyphenols, prevalent in plants and fungi, are investigated intensively in nutritional and clinical settings because of their beneficial bioactive properties. Due to their complexity, analysis with untargeted approaches is favorable, which typically use high-resolution mass spectrometry (HRMS) rather than low-resolution mass spectrometry (LRMS). Here, the advantages of HRMS were evaluated by thoroughly testing untargeted techniques and available online resources. By applying data-dependent acquisition on real-life urine samples, 27 features were annotated with spectral libraries, 88 with in silico fragmentation, and 113 by MS1 matching with PhytoHub, an online database containing >2000 polyphenols. Moreover, other exogenous and endogenous molecules were screened to measure chemical exposure and potential metabolic effects using the Exposome-Explorer database, further annotating 144 features. Additional polyphenol-related features were explored using various non-targeted analysis techniques including MassQL for glucuronide and sulfate neutral losses, and MetaboAnalyst for statistical analysis. As HRMS typically suffers a sensitivity loss compared to state-of-the-art LRMS used in targeted workflows, the gap between the two instrumental approaches was quantified in three spiked human matrices (urine, serum, plasma) as well as real-life urine samples. Both instruments showed feasible sensitivity, with median limits of detection in the spiked samples being 10-18 ng/mL for HRMS and 4.8-5.8 ng/mL for LRMS. The results demonstrate that, despite its intrinsic limitations, HRMS can readily be used for comprehensively investigating human polyphenol exposure. In the future, this work is expected to allow for linking human health effects with exposure patterns and toxicological mixture effects with other xenobiotics.

Synthesis, Characterization, and Antibacterial Activity of Ni-Substituted Krebs-type Sandwich-Tungstobismuthates Functionalized with Amino Acids.

Four new Ni-substituted Krebs-type sandwich-tungstobismuthates, K4Ni2[{Ni(ß-ala)(H2O)2}2{Ni(H2O)}2{Ni(H2O)(η2-ß-ala)}2(B-ß-BiW9O33)2]·49H2O {(ß-ala)4(Ni3)2(BiW9)2}, K3.5Na6.5[{Ni(η3-L-asp)}2(WO2)2(B-ß-BiW9O33)2]·36H2O·L-asp {(L-asp)2(NiW)2(BiW9)2}, K4Na6[{Ni(gly)(H2O)2}2(WO2)2(B-ß-BiW9O33)2]·86H2O {(gly)2(NiW)2(BiW9)2}, and K2Na8[{Ni(η2-serinol) (H2O)}2{Ni(H2O)2}2(B-ß-BiW9O33)2]·42H2O {(serinol)2Ni4(BiW9)2} have been synthesized by one-pot solution methods. All compounds have been characterized in the solid state by single-crystal X-ray diffraction (SXRD), powder X-ray diffraction (PXRD), elemental and thermogravimetric analyses, and infrared spectroscopy (IR), as well as by UV-vis spectroscopy in solution. The antibacterial activity of all compounds was studied against four bacterial strains by the determination of the minimum inhibitory concentration (MIC). The results showed that only {(ß-ala)4(Ni3)2(BiW9)2} demonstrates antibacterial activity (MIC is in the range from 8 to 256 µg/mL) compared to three other Ni-Krebs sandwiches.

Biochemical Investigations of Five Recombinantly Expressed Tyrosinases Reveal Two Novel Mechanisms Impacting Carbon Storage in Wetland Ecosystems.

Wetlands are globally distributed ecosystems characterized by predominantly anoxic soils, resulting from water-logging. Over the past millennia, low decomposition rates of organic matter led to the accumulation of 20-30% of the world's soil carbon pool in wetlands. Phenolic compounds are critically involved in stabilizing wetland carbon stores as they act as broad-scale inhibitors of hydrolytic enzymes. Tyrosinases are oxidoreductases capable of removing phenolic compounds in the presence of O2 by oxidizing them to the corresponding o-quinones. Herein, kinetic investigations (kcat and Km values) reveal that low-molecular-weight phenolic compounds naturally present within wetland ecosystems (including monophenols, diphenols, triphenols, and flavonoids) are accepted by five recombinantly expressed wetland tyrosinases (TYRs) as substrates. Investigations of the interactions between TYRs and wetland phenolics reveal two novel mechanisms that describe the global impact of TYRs on the wetland carbon cycle. First, it is shown that o-quinones (produced by TYRs from low-molecular-weight phenolic substrates) are capable of directly inactivating hydrolytic enzymes. Second, it is reported that o-quinones can interact with high-molecular-weight phenolic polymers (which inhibit hydrolytic enzymes) and remove them through precipitation. The balance between these two mechanisms will profoundly affect the fate of wetland carbon stocks, particularly in the wake of climate change.

The Novel Role of Tyrosinase Enzymes in the Storage of Globally Significant Amounts of Carbon in Wetland Ecosystems.

Over the last millennia, wetlands have been sequestering carbon from the atmosphere via photosynthesis at a higher rate than releasing it and, therefore, have globally accumulated 550 × 1015 g of carbon, which is equivalent to 73% of the atmospheric carbon pool. The accumulation of organic carbon in wetlands is effectuated by phenolic compounds, which suppress the degradation of soil organic matter by inhibiting the activity of organic-matter-degrading enzymes. The enzymatic removal of phenolic compounds by bacterial tyrosinases has historically been blocked by anoxic conditions in wetland soils, resulting from waterlogging. Bacterial tyrosinases are a subgroup of oxidoreductases that oxidatively remove phenolic compounds, coupled to the reduction of molecular oxygen to water. The biochemical properties of bacterial tyrosinases have been investigated thoroughly in vitro within recent decades, while investigations focused on carbon fluxes in wetlands on a macroscopic level have remained a thriving yet separated research area so far. In the wake of climate change, however, anoxic conditions in wetland soils are threatened by reduced rainfall and prolonged summer drought. This potentially allows tyrosinase enzymes to reduce the concentration of phenolic compounds, which in turn will increase the release of stored carbon back into the atmosphere. To offer compelling evidence for the novel concept that bacterial tyrosinases are among the key enzymes influencing carbon cycling in wetland ecosystems first, bacterial organisms indigenous to wetland ecosystems that harbor a TYR gene within their respective genome (tyr+) have been identified, which revealed a phylogenetically diverse community of tyr+ bacteria indigenous to wetlands based on genomic sequencing data. Bacterial TYR host organisms covering seven phyla (Acidobacteria, Actinobacteria, Bacteroidetes, Firmicutes, Nitrospirae, Planctomycetes, and Proteobacteria) have been identified within various wetland ecosystems (peatlands, marshes, mangrove forests, bogs, and alkaline soda lakes) which cover a climatic continuum ranging from high arctic to tropic ecosystems. Second, it is demonstrated that (in vitro) bacterial TYR activity is commonly observed at pH values characteristic for wetland ecosystems (ranging from pH 3.5 in peatlands and freshwater swamps to pH 9.0 in soda lakes and freshwater marshes) and toward phenolic compounds naturally present within wetland environments (p-coumaric acid, gallic acid, protocatechuic acid, p-hydroxybenzoic acid, caffeic acid, catechin, and epicatechin). Third, analyzing the available data confirmed that bacterial host organisms tend to exhibit in vitro growth optima at pH values similar to their respective wetland habitats. Based on these findings, it is concluded that, following increased aeration of previously anoxic wetland soils due to climate change, TYRs are among the enzymes capable of reducing the concentration of phenolic compounds present within wetland ecosystems, which will potentially destabilize vast amounts of carbon stored in these ecosystems. Finally, promising approaches to mitigate the detrimental effects of increased TYR activity in wetland ecosystems and the requirement of future investigations of the abundance and activity of TYRs in an environmental setting are presented.

Similar but Still Different: Which Amino Acid Residues Are Responsible for Varying Activities in Type-III Copper Enzymes?

Type-III copper enzymes like polyphenol oxidases (PPOs) are ubiquitous among organisms and play a significant role in the formation of pigments. PPOs comprise different enzyme groups, including tyrosinases (TYRs) and catechol oxidases (COs). TYRs catalyze the o-hydroxylation of monophenols and the oxidation of o-diphenols to the corresponding o-quinones (EC 1.14.18.1). In contrast, COs only catalyze the oxidation of o-diphenols to the corresponding o-quinones (EC 1.10.3.1). To date (August 2020), 102 PDB entries encompassing 18 different proteins from 16 organisms and several mutants have been reported, identifying key residues for tyrosinase activity. The structural similarity between TYRs and COs, especially within and around the active center, complicates the elucidation of their modes of action on a structural basis. However, mutagenesis studies illuminate residues that influence the two activities and show that crystallography on its own cannot elucidate the enzymatic activity mode. Several amino acid residues around the dicopper active center have been proposed to play an essential role in the two different activities. Herein, we critically review the role of all residues identified so far that putatively affect the two activities of PPOs.

Interweaving Disciplines to Advance Chemistry: Applying Polyoxometalates in Biology.

This Viewpoint brings awareness of the challenges and subsequent breakthroughs at the intersection of different disciplines, illustrated by the example of the influence biological entities exerted on a huge class of inorganic coordination compounds, called polyoxometalates (POMs). We highlight the possible effects of biological systems on POMs that need to be considered, thereby emphasizing the depth and complexity of interdisciplinary work. We map POMs' structural, electrochemical, and stability properties in the presence of biomolecules and stress the potential challenges related to inorganic coordination chemistry carried out in biological systems. This Viewpoint shows that new chemistry is available at the intersections between disciplines and aims to guide the community toward a discussion about current as well as future trends in truly interdisciplinary work.

Defect {(WVIO7)WVI4} and Full {(WVIO7)WVI5} Pentagonal Units as Synthons for the Generation of Nanosized Main Group V Heteropolyoxotungstates.

We report on the synthesis and characterization of three new nanosized main group V heteropolyoxotungstates KxNay[H2(XWVI9O33)(WVI5O12)(X2WVI29O103)]·nH2O {X3W43} (x = 11, y = 16, and n = 115.5 for X = SbIII; x = 20, y = 7, and n = 68 for X = BiIII) and K8Na15[H16(CoII(H2O)2)0.9(CoII(H2O)3)2(WVI3.1O14)(SbIIIWVI9O33)(SbIII2WVI30O106)(H2O)]·53H2O {Co3Sb3W42}. On the basis of the key parameters for the one-pot synthesis strategy of {Bi3W43}, a rational step-by-step approach was developed using the known Krebs-type polyoxotungstate (POT) K12[SbV2WVI22O74(OH)2]·27H2O {Sb2W22} as a nonlacunary precursor leading to the synthesis and characterization of {Sb3W43} and {Co3Sb3W42}. Solid-state characterization of the three new representatives {Bi3W43}, {Sb3W43}, and {Co3Sb3W42} by single-crystal and powder X-ray diffraction (XRD), IR spectroscopy, thermogravimetric analysis (TGA), energy-dispersive X-ray analysis (EDX), X-ray photoelectron spectroscopy (XPS), and elemental analysis, along with characterization in solution by UV/vis spectroscopy shows that {Bi3W43}, {Sb3W43}, and {Co3Sb3W42} represent the first main group V heteropolyoxotungstates encapsulating a defect {(WVIO7)WVI4} ({X3W43}, X = BiIII and SbIII) or full {(WVIO7)WVI5} ({Co3Sb3W42}) pentagonal unit. With 43 tungsten metal centers, {X3W43} (X = BiIII and SbIII) are the largest unsubstituted tungstoantimonate- and bismuthate clusters reported to date. By using time-dependent UV/vis spectroscopy, the isostructural representatives {Sb3W43} and {Bi3W43} were subjected to a comprehensive study on their catalytic properties as homogeneous electron-transfer catalysts for the reduction of K3[FeIII(CN)6] as a model substrate revealing up to 5.8 times higher substrate conversions in the first 240 min (35% for {Sb3W43}, 29% for {Bi3W43}) as compared to the uncatalyzed reaction (<6% without catalyst after 240 min) under otherwise identical conditions.

The Smallest Polyoxotungstate Retained by TRIS-Stabilization.

A polycondensation reaction of the orthotungstate anion WO42-, buffered at pH 7.5 in a TRIS-HCl (0.15 M) solution, results in the first example of a discrete polyoxotungstate anion, with just two W ions stabilized with TRIS ligands. It was isolated and characterized as Na2[WVI2O6(C4O3NH10)2]·6H2O by single-crystal and powder X-ray diffraction, FT-IR spectroscopy, thermogravimetrical analysis (TGA), and elemental analysis in solid state and by electro-spray ionization mass spectrometry (ESI-MS), 13C, and 183W NMR, as well as Raman spectroscopy in solution. This synthesis demonstrates the crucial and new role of the added tris-alkoxy ligand in the development of a new hybrid TRIS-isopolytungstate with the lowest known nuclearity (so far) and the terminal oxygens substituted with two nitrogen atoms arising from amines of the TRIS ligands.

Speciation of Transition-Metal-Substituted Keggin-Type Silicotungstates Affected by the Co-crystallization Conditions with Proteinase K.

We report on the synthesis of the tetrasubstituted sandwich-type Keggin silicotungstates as the pure Na salts Na14[(A-α-SiW10O37)2{Co4(OH)2(H2O)2}]·37H2O (Na{SiW10Co2}2) and Na14[(A-α-SiW10O37)2{Ni4(OH)2(H2O)2}]·77.5H2O (Na{SiW10Ni2}2), which were prepared by applying a new synthesis protocol and characterized thoroughly in the solid state by single-crystal and powder X-ray diffraction, IR spectroscopy, thermogravimetric analysis, and elemental analysis. Proteinase K was applied as a model protein and the polyoxotungstate (POT)-protein interactions of Na{SiW10Co2}2 and Na{SiW10Ni2}2 were studied side by side with the literature-known K5Na3[A-α-SiW9O34(OH)3{Co4(OAc)3}]·28.5H2O ({SiW9Co4}) featuring the same number of transition metals. Testing the solution behavior of applied POTs under the crystallization conditions (sodium acetate buffer, pH 5.5) by time-dependent UV/vis spectroscopy and electrospray ionization mass spectrometry speciation studies revealed an initial dissociation of the sandwich POTs to the disubstituted Keggin anions HxNa5-x[SiW10Co2O38]3- and HxNa5-x[SiW10Ni2O38]3- ({SiW10M2}, M = CoII and NiII) followed by partial rearrangement to the monosubstituted compounds (α-{SiW11Co} and α-{SiW11Ni}) after 1 week of aging. The protein crystal structure analysis revealed monosubstituted α-Keggin POTs in two conserved binding positions for all three investigated compounds, with one of these positions featuring a covalent attachment of the POT anion to an aspartate carboxylate. Despite the presence of both mono- and disubstituted anions in a crystallization mixture, proteinase K selectively binds to monosubstituted anions because of their preferred charge density for POT-protein interaction.

Aluminum-Substituted Keggin Germanotungstate [HAl(H2O)GeW11O39]4-: Synthesis, Characterization, and Antibacterial Activity.

We report on the new monosubstituted aluminum Keggin-type germanotungstate (C4H12N)4[HAlGeW11O39(H2O)]·11H2O ([Al(H2O)GeW11]4-), which has been synthesized at room temperature via rearrangement of the dilacunary [γ-GeW10O36]8- polyoxometalate precursor. [Al(H2O)GeW11]4- has been characterized thoroughly both in the solid state by single-crystal and powder X-ray diffraction, IR spectroscopy, thermogravimetric analysis, and elemental analysis as well as in solution by cyclic voltammetry (CV) 183W, 27Al NMR and UV-vis spectroscopy. A study on the antibacterial properties of [Al(H2O)GeW11]4- and the known aluminum(III)-centered Keggin polyoxotungstates (Al-POTs) α-Na5[AlW12O40] (α-[AlW12O40]5-) and Na6[Al(AlOH2)W11O39] ([Al(AlOH2)W11O39]6-) revealed enhanced activity for all three Al-POTs against the Gram-negative bacterium Moraxella catarrhalis (minimum inhibitory concentration (MIC) up to 4 µg mL-1) and the Gram-positive Enterococcus faecalis (MIC up to 128 µg mL-1) compared to the inactive Al(NO3)3 salt (MIC > 256 µg mL-1). CV indicates the redox activity of the Al-POTs as a dominating factor for the observed antibacterial activity with increased tendency to reduction, resulting in increased antibacterial activity of the POT.

Expression, Purification, and Characterization of a Well-Adapted Tyrosinase from Peatlands Identified by Partial Community Analysis.

In peatlands, bacterial tyrosinases (TYRs) are proposed to act as key regulators of carbon storage by removing phenolic compounds, which inhibit the degradation of organic carbon. Historically, TYR activity has been blocked by anoxia resulting from persistent waterlogging; however, recent events of prolonged summer drought have boosted TYR activity and, consequently, the release of carbon stored in the form of organic compounds from peatlands. Since 30% of the global soil carbon stock is stored in peatlands, a profound understanding of the production and activity of TYRs is essential to assess the impact of carbon dioxide emitted from peatlands on climate change. TYR partial sequences identified by degenerated primers suggest a versatile TYR enzyme community naturally present in peatlands, which is produced by a phylogenetically diverse spectrum of bacteria, including Proteobacteria and Actinobacteria. One full-length sequence of an extracellular TYR (SzTYR) identified from a soda-rich inland salt marsh has been heterologously expressed and purified. SzTYR exhibits a molecular mass of 30 891.8 Da and shows a pH optimum of 9.0. Spectroscopic studies and kinetic investigations characterized SzTYR as a tyrosinase and proved its activity toward monophenols (coumaric acid), diphenols (caffeic acid, protocatechuic acid), and triphenols (gallic acid) naturally present in peatlands.

Polyoxometalates in solution: speciation under spotlight.

Polyoxometalates (POMs) are a large group of anionic polynuclear metal-oxo clusters with discrete and chemically modifiable structures. In most aqueous POM solutions, numerous, and often highly negatively charged, species of different nuclearities are formed. It is rather difficult to determine the dominant POM species or their combination, which is responsible for the specific POM activity, during a particular application. Thus, the identification of all individual speciation profiles is essential for the successful implementation of POMs in solution applications. This review article summarizes species that are present in isopoly- and heteropolyvanadates, -niobates, -molybdates and -tungstates aqueous solutions and covers their stability and transformations. The ion-distribution diagrams over a wide pH range are presented in a comprehensive manner. These diagrams are intended for the targeted use of POMs, and in a clear form shows species that are in equilibrium at the given pH value. Thus, the data accumulated in this review can serve as both a starting point and a complete reference material for determining the composition of POM solutions. Some examples are highlighted where the POM speciation studies led to a detailed understanding of their role in applications. In doing so, we aim to motivate the POM community for more speciation studies and to make the subject more comprehensible, both for synthetic POM chemists and for scientists with different backgrounds interested in applying POMs in biological, medical, electrochemical, supramolecular and nanochemistry fields, or as homogeneous catalysts and other water-soluble materials.

Incorporation of CrIII into a Keggin Polyoxometalate as a Chemical Strategy to Stabilize a Labile {CrIIIO4} Tetrahedral Conformation and Promote Unattended Single-Ion Magnet Properties.

Polyoxometalates (POMs) provide rigid and highly symmetric coordination sites and can be used as a strategy for the stabilization of magnetic ions. Herein, we report a new member of the Keggin archetype, the Cr-centered Keggin anion [α-CrW12O40]5- (CrW12), with the unusual tetrahedral coordination of CrIII reported for the first time in POMs conferring unattended magnetic properties. POM chemistry has recently presented excellent examples of single-molecule and single-ion magnets (SMMs and SIMs) as well as molecular spin qubits; however, the majority of POM-based SIMs reported to date contain lanthanoid ions. CrW12, as the first example of a chromium(III) SIM, exhibits slow relaxation of magnetization and quantum tunneling with a single-ion magnetic behavior even above 10 K with an energy barrier for the reversal of the magnetization of 3.0 K. The first 3d-metal SIM based on a nonlacunary Keggin anion is the foundation for a new research area in POM chemistry.

Binding of a Fatty Acid-Functionalized Anderson-Type Polyoxometalate to Human Serum Albumin.

The Anderson-type hexamolybdoaluminate functionalized with lauric acid (LA), (TBA)3[Al(OH)3Mo6O18{(OCH2)3CNHCOC11H23}]·9H2O (TBA-AlMo6-LA, where TBA = tetrabutylammonium), was prepared via two synthetic routes and characterized by thermogravimetric and elemental analyses, mass spectrometry, IR and 1H NMR spectroscopy, and powder and single-crystal X-ray diffraction. The interaction of TBA-AlMo6-LA with human serum albumin (HSA) was investigated via fluorescence and circular dichroism spectroscopy. The results revealed that TBA-AlMo6-LA binds strongly to HSA (63% quenching at an HSA/TBA-AlMo6-LA ratio of 1:1), exhibiting static quenching. In contrast to TBA-AlMo6-LA, the nonfunctionalized polyoxometalate, Na3(H2O)6[Al(OH)6Mo6O18]·2H2O (AlMo6), showed weak binding toward HSA (22% quenching at a HSA/AlMo6 ratio of 1:25). HSA binding was confirmed by X-ray structure analysis of the HSA-Myr-AlMo6-LA complex (Myr = myristate). These results provide a promising lead for the design of novel polyoxometalate-based hybrids that are able to exploit HSA as a delivery vehicle to improve their pharmacokinetics and bioactivity.

Synthesis, Characterization, and Phosphoesterase Activity of a Series of 4f- and 4d-Sandwich-Type Germanotungstates [(n-C4H9)4N]l/mH2[(M(H2O)3)(γ-GeW10O35)2] (M = CeIII, NdIII, GdIII, ErIII, l = 7; ZrIV, m = 6).

We report on a family of five new 4f- and 4d-doped sandwich-type germanotungstates with the general formula [(n-C4H9)4N]l/mH2[(M(H2O)3)(γ-GeW10O35)2]·3(CH3)2CO [M(H2O)3(GeW10)2] (M = CeIII, NdIII, GdIII, ErIII, l = 7; ZrIV, m = 6), which have been synthesized at room temperature in an acetone-water mixture. Among the compound series, [Zr(H2O)3(GeW10)2]8-, which has been obtained in the presence of 30% H2O2, represents the first example of a 4d-substituted germanotungstate incorporating the intact dilacunary [γ-GeIVW10O36]8- building block. All compounds were characterized thoroughly in the solid state by single-crystal and powder X-ray diffraction (XRD), IR spectroscopy, thermogravimetric analysis (TGA), and elemental analysis and in solution by NMR and UV-vis spectroscopy. The phosphoesterase activity of [Ce(H2O)3(GeW10)2]9- and [Zr(H2O)3(GeW10)2]8- toward the model substrates 4-nitrophenyl phosphate (NPP) and O,O-dimethyl O-(4-nitrophenyl) phosphate (DMNP) was monitored with 1H- and 31P-NMR spectroscopy revealing an acceleration of the hydrolytic reaction by an order of magnitude (kcorr = 3.44 (±0.30) × 10-4 min-1 for [Ce(H2O)3(GeW10)2]9- and kcorr = 5.36 (±0.05) × 10-4 min-1 for [Zr(H2O)3(GeW10)2]8-) as compared to the uncatalyzed reaction (kuncat = 2.60 (±0.10) × 10-5 min-1). [Ce(H2O)3(GeW10)2]9- demonstrated improved antibacterial activity toward Moraxella catarrhalis (MIC 32 µg/mL), compared to the unsubstituted [GeW10O36]8- POM (MIC 64 µg/mL).

Cation-Directed Synthetic Strategy Using 4f Tungstoantimonates as Nonlacunary Precursors for the Generation of 3d-4f Clusters.

The first synthetic pathway using a series of four nonlacunary 4f-heterometal-substituted polyoxotungstate clusters Na21[(Ln(H2O)(OH)2(CH3COO))3(WO4)(SbW9O33)3]·nH2O (NaLnSbW9; Ln = TbIII, DyIII, HoIII, ErIII, YIII) as precursors for the directed preparation of nine new 3d-4f heterometallic tungstoantimonates K5Na12H3[TM(H2O)Ln3(H2O)5(W3O11)(SbW9O33)3]·nH2O (KTMLnSbW9; TM = CoII, NiII; Ln = TbIII, DyIII, HoIII, ErIII, YIII) has been developed. Systematic studies revealed an increased K content in the aqueous acidic reaction mixture to be the key step in the cation-directed preparation of 3d-4f compounds; among those, the Co-containing members represent the first examples of KCoLnSbW9 (Ln = TbIII, DyIII, HoIII, ErIII, YIII) heterometallic tungstoantimonates exhibiting the SbW9 building block. All 13 compounds have been characterized thoroughly in the solid state by powder and single-crystal X-ray diffraction (XRD), revealing a cyclic trimeric polyoxometalate architecture with three SbW9 units encapsulating a planar triangle of LnIII ions in the case of NaLnSbW9 and a heterometallic core of one TMII and three LnIII for KTMLnSbW9 (TM = CoII, NiII; Ln = TbIII, DyIII, HoIII, ErIII, YIII). The results obtained by XRD are supplemented by complementary characterization methods in the solid state such as IR spectroscopy, thermogravimetric analysis, and elemental analysis as well as in solution by UV-vis spectroscopy. Detailed magnetic studies on the representative compounds KTMDySbW9 (TM = CoII, NiII) and KCoYSbW9 of the series revealed field-induced slow magnetic relaxation.

The Aquaporin-3-Inhibiting Potential of Polyoxotungstates.

Polyoxometalates (POMs) are of increasing interest due to their proven anticancer activities. Aquaporins (AQPs) were found to be overexpressed in tumors bringing particular attention to their inhibitors as anticancer drugs. Herein, we report for the first time the ability of polyoxotungstates (POTs), such as of Wells-Dawson P2W18, P2W12, and P2W15, and Preyssler P5W30 structures, to affect aquaporin-3 (AQP3) activity and impair melanoma cell migration. The tested POTs were revealed to inhibit AQP3 function with different effects, with P2W18, P2W12, and P5W30 being the most potent (50% inhibitory concentration (IC50) = 0.8, 2.8, and 3.2 µM), and P2W15 being the weakest (IC50 > 100 µM). The selectivity of P2W18 toward AQP3 was confirmed in yeast cells transformed with human aquaglyceroporins. The effect of P2W12 and P2W18 on melanoma cells that highly express AQP3 revealed an impairment of cell migration between 55% and 65% after 24 h, indicating that the anticancer properties of these compounds may in part be due to the blockage of AQP3-mediated permeability. Altogether, our data revealed that P2W18 strongly affects AQP3 activity and cancer cell growth, unveiling its potential as an anticancer drug against tumors where AQP3 is highly expressed.

Identification of Amino Acid Residues Responsible for C-H Activation in Type-III Copper Enzymes by Generating Tyrosinase Activity in a Catechol Oxidase.

Tyrosinases (TYRs) catalyze the hydroxylation of phenols and the oxidation of the resulting o-diphenols to o-quinones, while catechol oxidases (COs) exhibit only the latter activity. Aurone synthase (AUS) is not able to react with classical tyrosinase substrates, such as tyramine and l-tyrosine, while it can hydroxylate its natural substrate isoliquiritigenin. The structural difference of TYRs, COs, and AUS at the heart of their divergent catalytic activities is still a puzzle. Therefore, a library of 39 mutants of AUS from Coreopsis grandiflora (CgAUS) was generated and the activity studies showed that the reactivity of the three conserved histidines (HisA2 , HisB1 , and HisB2 ) is tuned by their adjacent residues (HisB1 +1, HisB2 +1, and waterkeeper residue) either to react as stronger bases or / and to stabilize a position permissive for substrate proton shuffling. This provides the understanding for C-H activation based on the type-III copper center to be used in future biotechnological processes.