Metal-binding Ability Research Articles

Effectiveness and Mechanisms of Recoverable Magnetic Nanoparticles on Mitigating Golden Mussel Biofouling.

Mussel biofouling has become a problem in aquatic ecosystems, causing significant ecological impact and huge economic loss globally. Although several strategies have been proposed and tested, efficient and environment-friendly antifouling methods are still scarce. Here, we investigated the effects of recoverable magnetic ferroferric oxide nanoparticles (Fe3O4-NPs) with different sizes (10 and 100 nm), coatings (polyethylene glycol and polylysine), and concentrations (0.01 and 0.1 mg/L) on byssus adhesion-mediated biofouling by the notorious golden mussel Limnoperna fortunei. The results showed that magnetic Fe3O4-NPs, especially negatively charged polyethylene glycol-coated Fe3O4-NPs, size- and concentration-dependently reduced the byssus production, performance (breaking force and failure location), and adhesion rate. Further investigations on mechanisms showed that the down-regulation of foot protein 2 (Lffp-2) and energy-related metabolic pathways inhibited byssus production. The declined gene expression level and metal-binding ability of Lffp-2 significantly affected foot protein interactions, further reducing the plaque size and byssus performance. In addition, the change in the water redox state likely reduced byssus performance by preventing the interface interactions between the substrate and foot proteins. Our results confirm the effectiveness and underlying mechanisms of magnetic Fe3O4-NPs on mitigating L. fortunei biofouling, thus providing a reference for developing efficient and environment-friendly antifouling strategies against fouling mussels.

Nitric oxide-donating and reactive oxygen species-responsive prochelators based on 8-hydroxyquinoline as anticancer agents.

Metal ion chelators based on 8-hydroxyquinoline (8-HQ) have been widely explored for the treatment of many diseases. When aimed at being developed into potent anticancer agent, a largely unmet issue is how to avoid nonspecific chelation of metal ions by 8-HQ in normal cells or tissues. In the current work, a two-step strategy was employed to both enhance the anticancer activity of 8-HQ and improve its cancer cell specificity. Considering the well-known anticancer activity of nitric oxide (NO), NO donor furoxan was first connected to 8-HQ to construct HQ-NO conjugates. These conjugates were screened for their cytotoxicity, metal-binding ability, and NO-releasing efficiency. Selected conjugates were further modified with a ROS-responsive moiety to afford prochelators. Among all the target compounds, prodrug HQ-NO-11 was found to potently inhibit the proliferation of many cancer cells but not normal cells. The abilities of metal chelation and NO generation by HQ-NO-11 were confirmed by various methods and were demonstrated to be essential for the anticancer activity of HQ-NO-11. Invivo studies revealed that HQ-NO-11 inhibited the growth of SW1990 xenograft to a larger extent than 8-HQ. Our results showcase a general method for designing novel 8-HQ derivatives and shed light on obtaining more controllable metal chelators.

Mediation of metal chelation in cysteine-derived tetramate systems.

A study of bicyclic tetramates modified with a bulky ester, which leads to steric hindrance of distal chelating atoms as a route for the alteration of metal binding ability is reported. This approach required the development of a direct method for the synthesis of different esters of cysteine from cystine, which then provided access to bicyclic tetramates by Dieckmann cyclisation. Further derivation to ketones and carboxamides by Grignard addition and transamination reactions respectively provided rapid access to a chemical library of tetramates with diverse substitution. Of interest is that bicyclic tetramate ketones and carboxamides showed different tautomeric and metal binding behaviour in solution. Significantly, in both systems, the incorporation of bulky C-5 esters at the bridging position not only reduced metal binding, but also enhanced antibacterial potencies against Gram-positive MRSA bacteria. Those tetramates with antibacterial activity which was not metal dependent showed physiochemical properties of MSA of 559–737 Å2, MW of 427–577 Da, clogP of 1.8–6.1, clogD7.4 of −1.7 to 3.7, PSA of 83–109 Å2 and relative PSA of 12–15% and were generally Lipinski rule compliant. A subset of tetramates exhibited good selectivity towards prokaryotic bacterial cells. Given that the work reported herein is synthesis-led, without the underpinning detailed mechanistic understanding of biological/biochemical mechanism, that the most active compounds occupy a small region of chemical space as defined by MW, clogP, PSA and %PSA is of interest. Overall, the bicyclic tetramate template is a promising structural motif for the development of novel antibacterial drugs, with good anti-MRSA potencies and appropriate drug-like physiochemical properties, coupled with a potential for multi-targeting mechanisms and low eukaryotic cytotoxicity.

Gravity-driven multifunctional microporous membranes for household water treatment: Simultaneous pathogenic disinfection, metal recycling, and biofouling mitigation

In remote industrial areas, access to safe drinking water is often hindered by local water contamination and unreliable infrastructure. Hence, energy-effective water treatment and safe storage technology is vital and highly desirable to tackle complex water pollution caused by heavy metals and pathogenic contaminants for boundary circumstances. Here, we originally engineered a multifunctional gravity-driven membrane system to provide a household-level approach to pathogenic disinfection and biofouling prevention. Facile one-step polyphenol deposition strategy was applied on microporous membranes, tailoring the membrane surface with enhanced positive charge, favorable hydrophilicity and metal-binding ability. When applied in gravity-driven filtration of simulated multi-component wastewater containing heavy metals and pathogenic contaminants, the as-prepared membrane achieved >6 log pathogenic reduction. Metal ions could be in-situ captured and stabilized at membrane interface via natural binding and reduction by polyphenol-engineered surface chemistry, endowing ensuing membranes with improved antibacterial activity. By virtue of enhanced membrane surface properties and antimicrobial synergy, such engineered membrane platform showed potent efficacy in biofilm prevention with97% reduced ATP content and easy permeation recovery (>90%) in cyclic operation. This study is promising for developing low-energy and easy-to-use membrane system to improve contaminated water sources in remote and underdeveloped areas at household level.

Two Unconventional Metallothioneins in the Apple Snail Pomacea bridgesii Have Lost Their Metal Specificity during Adaptation to Freshwater Habitats.

Metallothioneins (MTs) are a diverse group of proteins responsible for the control of metal homeostasis and detoxification. To investigate the impact that environmental conditions might have had on the metal-binding abilities of these proteins, we have characterized the MTs from the apple snail Pomacea bridgesii, a gastropod species belonging to the class of Caenogastropoda with an amphibious lifestyle facing diverse situations of metal bioavailability. P. bridgesii has two structurally divergent MTs, named PbrMT1 and PbrMT2, that are longer than other gastropod MTs due to the presence of extra sequence motifs and metal-binding domains. We have characterized the Zn(II), Cd(II), and Cu(I) binding abilities of these two MTs after their heterologous expression in E. coli. Our results have revealed that despite their structural differences, both MTs share an unspecific metal-binding character, and a great ability to cope with elevated amounts of different metal ions. Our analyses have also revealed slight divergences in their metal-binding features: PbrMT1 shows a more pronounced Zn(II)-thionein character than PbrMT2, while the latter has a stronger Cu(I)-thionein character. The characterization of these two unconventional PbrMTs supports the loss of the metal-binding specificity during the evolution of the MTs of the Ampullariid family, and further suggests an evolutionary link of this loss with the adaptation of these gastropod lineages to metal-poor freshwater habitats.

Obtaining chitosan polysaccharide from various natural raw materials and studying its metal-binding ability

The article considers the structure and properties of chitosan obtained from natural sources of plant and animal origin. The ability of chitosan to complex formation with copper ions is studied. The isotherm of ion sorption is given, and the quantitative values of the complexation process are calculated according to the classical theory and the theory of RCC.

Metal-binding ability of sodium alginate relative to Fe3+ and Cu2+ ions

The article substantiates and experimentally confirms the possibility of a non-starch polysaccharide – sodium alginate to form complex compounds with iron (III) and copper (II) ions. Alginates are used in medical practice as an enterosorbent for severe poisoning with metal ions due to their ability to form complexes with heavy metals, insoluble in water.

Nanomagnetite and Chitosan Affect Seed Germination and Growth of Wheat (Triticum Aestivum L.)

Iron deficiency is one of the limiting factors of plant growth. In recent years nanotechnology has attracted attention to producing forms of iron that are available to plants. Chitosan, which is widely used in agriculture due to its unique properties, has metal-binding abilities. We decided to study nanomagnetite and chitosan's combined action to improve seed germination and growth of wheat (Triticum aestivum L.). The thumbnail method was used in the study. The seeds were treated with magnetite suspension and chitosan solutions before planting. An increase in plant growth, root, and shoot length of plants was found in all seed treatment methods (chitosan, nanomagnetite, and combined action). This increase was significantly dependent on the concentration of chitosan. However, chitosan and magnetite combination was 1.3 times more effective for seed germination than pure chitosan or magnetite. A mechanism based on chitosan's ability to promote the better transfer of iron ions to plants is proposed. However, further investigation for the optimization of chitosan/magnetite proportions for improving seed growth is required.

Dynamic evolution of humic acids during anaerobic digestion: Exploring an effective auxiliary agent for heavy metal remediation

Information on the dynamic evolution of humic acid (HA) from anaerobic digestate and the potential of HA serving as an effective agent for remedying heavy metals is rather scarce. This study monitored the evolution of the structure and functional groups and metal-binding abilities of HA during chicken manure and corn stover anaerobic digestion (AD) processes. Higher increases in aromatic (41–66%) and oxygen-containing functional groups (37–45%) were observed in HA from the AD of corn stover, resulting in higher metal-binding abilities for Cu(II), Co(II), and Ni(II) than those of chicken manure AD. Moreover, HA extracted from fast (before day 12 for chicken manure and day 16 for corn stover), and slow (day 40) methane production stages performed different complexation capacities for the heavy metals. These results reveal the mechanisms of HA and heavy metal interactions, and confirm the potential of HA extracted from AD process for the remediation of heavy metals.

Two metallothionein genes highly expressed in rice nodes are involved in distribution of Zn to the grain.

A rice node is a hub for distribution of mineral elements; however, most genes highly expressed in the node have not been functionally characterized. Transcriptomic analysis of a rice node revealed that two metallothionein genes, OsMT2b and OsMT2c, were highly expressed in the node I. We functionally characterized these genes in terms of gene expression pattern, cellular and subcellular localization, phenotypic analysis of the single and double knockout mutants and metal-binding ability. Both OsMT2b and OsMT2c were mainly and constitutively expressed in the phloem region of enlarged and diffuse vascular bundles in the nodes and of the anther. Knockout of either OsMT2b or OsMT2c increased zinc (Zn) accumulation in the nodes, but decreased Zn distribution to the panicle, resulting in decreased grain yield. A double mutant, osmt2bmt2c, showed further negative effects on the Zn distribution and grain yield. By contrast, knockout of OsMT2b had a small effect on copper (Cu) accumulation. Both OsMT2b and OsMT2c showed binding ability with Zn, whereas only OsMT2b showed binding ability with Cu in yeast. Our results suggest that both OsMT2b and OsMT2c play an important role mainly in the distribution of Zn to grain through chelation and subsequent transport of Zn in the phloem in rice.

Thermal characteristics and cadmium binding behavior of EC-ELP fusion polypeptides

Elastin-like polypeptides (ELPs) are stimulus–responsive protein-based biopolymers that exhibit phase transition behavior. By joining them to synthetic phytochelatin (EC), EC–ELP fusion proteins with temperature sensitivity and metal-binding functionality were generated to remove heavy metal ions biologically. Three different EC domains (EC10, EC20, EC30) were incorporated into the ELP, and the EC–ELP fusion proteins were expressed in E. coli. Their thermal properties and metal binding abilities were then investigated according to the EC length. In addition, the feasibility of reusing EC–ELPs and the cadmium ion binding affinity of reused EC–ELPs were explored.

Metal binding ability of microbial natural metal chelators and potential applications

Covering: up to 2019 Siderophores are natural products which have attracted interest due to their ability to bind metal ions with high affinities. This review considers the complex formation of these natural chelators with all kinds of different metal and metalloid ions and links their chelating properties to potential biotechnological applications and towards environmental context.

Modular quadruplex-duplex hybrids as biomolecular scaffolds for asymmetric Michael addition reactions.

Asymmetric synthesis based on DNA scaffolds has been actively exploited because of the advantages of DNA such as diverse tertiary structures, chemical stability, and easy handling. Since duplex DNA-based hybrid catalysts have demonstrated this remarkable capability, efforts have been made to investigate new biomolecular scaffolds. Herein, we report modular quadruplex-duplex (QD) hybrid DNA catalysts containing bipyridine ligands and hydrogen donor moieties. The conformation, thermal stability, and metal-binding ability of modified QD hybrid DNA were characterized using spectroscopy. The QD hybrid-based DNA catalysts were successfully applied to asymmetric Michael addition reactions (86% conversion and 76% ee). This study describes a new type of DNA hybrid catalyst produced by the construction of a cooperative active site with a Lewis acid and a H-bond donor.

Complex formation processes and metal ion catalyzed oxidation of model peptides related to the metal binding site of the human prion protein

Interaction of copper(II) and nickel(II) ions with the Ac-PHAAAGTHSMKHM-NH2 tridecapeptide containing the His85, His96 and His111 binding sites of human prion protein has been studied by various techniques. pH-potentiometry, UV–Vis and circular dichroism spectroscopy were applied to study the stoichiometry, stability and structure of the copper(II) and nickel(II) complexes, while HPLC-MS and MS/MS were used for identifying the products of copper(II) catalyzed oxidation. The copper binding ability of shorter fragments, namely the nonapeptide Ac-PHAAAGTHS-NH2 and pentapeptide Ac-PHAAA-NH2 have also been studied. The tridecapeptide is able to bind three equivalent of copper(II) ion, since the histidine residues behave as independent metal binding sites. Nevertheless, the metal binding ability of histidine residue mimicking the octarepeat domain (His85) is decreased, while the other parts of the peptide mimicking the histidines outside the octarepeat domain bind the copper ions in comparable concentration. On the other hand, this peptide is able to coordinate only two equivalents of nickel ion on the domains outside the octarepeat region. Furthermore the His96 binding site is more effective for the nickel ions. Both histidine and methionine residues are sensitive for oxidation, the oxidation of these residues are proved, and in the case of the histidine residues follows the order His96 > His85 ≫ His111.

Clioquinol induces S-phase cell cycle arrest through the elevation of the calcium level in human neurotypic SH-SY5Y cells.

Clioquinol is recently considered to be the most promising drug for treating cancer and neurodegenerative diseases. However, its mode of action varies from different disease models. In this study, we found that clioquinol inhibited cell growth in human neurotypic SHSY-5Y cells, which was attributed to both S-phase cell-cycle arrest and autophagic cell death. Clioquinol increased the intracellular contents of iron and zinc as well as calcium as measured by ICP-AES. Staining of Fluo-3 confirmed an increase in the level of calcium. Analysis of the metal-binding ability of clioquinol showed that it was not a chelating agent of calcium ions and the elevation of intracellular calcium content is not achieved by clioquinol as an ionophore. CaCl2 could simulate or even aggravate the cytotoxicity of clioquinol and it increased S-phase cell cycle arrest induced by clioquinol in a concentration dependent manner. Staining of acridine orange demonstrated that autophagy induced by clioquinol was not affected by addition of calcium ions. In contrast, the intracellular calcium ion chelator BAPTA-am abolished the clioquinol-induced S phase arrest and reduced the cell death caused by clioquinol. The WB assay of cell cycle-related proteins (CDK2, p21 and p27) further confirmed that S phase arrest is positively correlated with intracellular calcium elevation, which was due to the alterations of the mRNA and protein levels of calcium pumps (SERCA and SPCA). Taken together, these data indicate that clioquinol regulates the level of intracellular calcium ions to induce S-phase cell cycle arrest in human SH-SY5Y cells. Our results demonstrate for the first time that an increase of intracellular calcium content is one of the mechanisms of clioquinol in the inhibition of human neurotypic SHSY-5Y cells.

The Enhancement of Metal-Binding Properties in Hemoglobin: The Role of Mild Damaging Factors.

X-ray studies revealed the considerable enhancement of metal-binding properties in human hemoglobin under exposure to mild damaging factors (in the presence of 0.09 M urea or upon heating for 30 min at 50 °C). Changes in the element composition of the hemoglobin monolayer, formed on the water subphase in the Langmuir trough, have been monitored in real time by the total external reflection X-ray fluorescence measurements. X-ray absorption spectroscopy has been applied to study the local environment of zinc ions bound on hemoglobin molecules. According to these data, each zinc ion is coordinated by four ligands, two of which are cysteine and histidine. The oxidative stress has been found to accelerate extensively the enhancement of metal-binding ability in protein. A two-stage mechanism has been proposed as a possible explanation of the observed phenomenon: First, in the presence of the mild damaging agents, protein molecules can undergo a transition from the native conformation to a more labile intermediate state that increases the accessibility of amino acid residues (in particular cysteine). At the second stage, oxidation of cysteine and the subsequent activation of cysteine SH groups can affect markedly the protein-metal interaction. The presented investigations provide a deeper insight into the pathogenesis of metabolic disorders that excessive concentrations of the endogenic toxicants might trigger in an organism.

Tandem oligomeric expression of metallothionein enhance heavy metal tolerance and bioaccumulation in Escherichia coli

Metallothioneins (MTs) are a family of low molecular weight, cysteine-rich, metal-binding proteins, which play important roles in metal homeostasis and heavy metal detoxification. In our previous study, a novel full length MT cDNA was successfully cloned from the freshwater crab (Sinopotamon henanense). In the present study, tandem repeats of two and three copies of the crab MT gene were integrated by overlap extension PCR (SOE-PCR) and expressed in Escherichia coli. The SUMO fusion expression system was adopted to increase the stability and solubility of the recombinant MT proteins. The recombinant proteins were purified and their metal-binding abilities were further analyzed by the ultraviolet absorption spectral scan. Furthermore, the metal tolerance and bioaccumulation of E. coli cells expressing oligomeric MTs were determined. Results showed that the recombinant plasmids pET28a-SUMO-2MT and pET28a-SUMO-3MT were successfully constructed. SDS-PAGE analysis showed that the SUMO-2MT and SUMO-3MT were expressed mainly in the soluble forms. Oligomeric MTs expression significantly enhanced Cu, Cd or Zn tolerance and accumulation in E. coli in the order: SUMO-3MT˃SUMO-2MT˃SUMO-MT˃control. Cells harboring pET28a-SUMO -3MT exhibited the highest Cu, Cd or Zn bioaccumulation at 5.8-fold, 3.1-fold or 6.7-fold higher than that of the control cells. Our research could lay a foundation for large-scale preparation of MTs and provide a scientific basis for bioremediation of heavy metal pollution by oligomeric MTs.

Potential metal-binding ability of proteins in the extracellular slime of Laccaria bicolor exposed to excessive Cu and Cd.

Ectomycorrhizal fungi can enhance the tolerance of plants to heavy metal stress by reducing the accumulation of heavy metals in the aerial parts of the plants. Extracellular chelation is a major mechanism of heavy metal tolerance in ectomycorrhizal fungi in which extracellular slime plays a fundamental role. The objectives of this study were to investigate the potential metal-binding ability and the protein composition of extracellular slime. The extracellular slime of Laccaria bicolor (L. bicolor) cultivated under Cd2+ and Cu2+ stress was separated using various ultrasonic pre-treatments. The protein content, composition, and metal content of the extracellular slime were measured. The results showed that the protein content in the extracellular slime significantly increased under both Cd2+ and Cu2+ stress. The SDS-PAGE profile showed that Cd2+ and Cu2+ stress induced the expression of several new proteins. Heavy metal quantification revealed that the Cd content fixed in the extracellular slime accounted for 22-28% of the metal fixed by the fungal mycelia. Meanwhile, no Cu was detected in the fungal extracellular slime, implying that the extracellular slime may not be effective for the fixation of essential metallic elements such as Cu. Taken together, these results provided evidence that L. bicolor was able to ameliorate the intracellular Cd content by stimulating extracellular slime exudation and altering the composition of the proteins therein. Nevertheless, this blocking strategy may be effective only for the non-essential element Cd and was ineffective for the physiological element Cu.

Dominin and Segon Form Multiprotein Particles in the Plasma of Eastern Oysters (Crassostrea virginica) and Are Likely Involved in Shell Formation.

Dominin and segon are two proteins purified and characterized from the plasma of eastern oysters Crassostrea virginica, making up about 70% of the total plasma proteins. Their proposed functions are in host defense based on their pathogen binding properties and in metal metabolism based on their metal binding abilities. In the present study, the two proteins were further studied for their native states in circulation and extrapallial fluid and their possible involvement in shell formation. Two-dimensional electrophoresis confirmed that the oyster plasma was dominated by a few major proteins and size exclusion chromatography indicated that these proteins were present in circulation in a morphologically homogenous form. Density gradient ultracentrifugation in Cesium Chloride isolated morphologically homogenous particles of about 25 nm in diameter from the plasma and extrapallial fluids. Polyacrylamide gel electrophoresis identified dominin, segon and an unidentified protein as the principal components of the particles and the three proteins likely formed a multiprotein complex that associated to form the particle. Additionally, three major proteins extracted from shell organic matrix were identified based on the apparent molecular weight in SDS-PAGE to correspond to the three major proteins of plasma and protein particles. Moreover, the hemocyte expression of dominin and segon genes measured by real-time RT-PCR increased significantly upon the initiation of shell repair and were significantly greater in younger oysters. These findings suggest that dominin and segon form protein particles by association with each other and perhaps some other major plasma proteins and play a significant role in oyster shell formation.

Functional Analysis and Characterization of the Metal Bound Sco Protein from Thermus thermophilus

Cytochrome c oxidase (CcO) is the final complex of the respiratory electron transport chain, which is a set of complexes that participate in the production of adenosine triphosphate (ATP), the main form of energy used by cells. CuA is the initial electron acceptor of CcO and consists of two copper ions bridged by two cysteines and ligated by two histidines, a methionine and the carbonyl backbone of a nearby glutamine. Sco proteins have been implicated in the assembly of the CuA, site but the exact role that it plays is the subject of much debate and study. Previous work has found that Sco proteins share a fold similar to thioredoxins, pointing toward the possibility of thiol‐disulfide oxidoreductase activity as a function of the protein. The ability of Thermus thermophilus Sco (TtSco) to act as a thiol‐disulfide oxidoreductase has been probed using an experiment to trap the protein‐protein intermediate between TtSco and its target, Thermus thermophilus CuA (TtCuA). The effect of nearby amino acids and metal binding prior to reaction have been determined for the formation of the protein‐protein intermediate, the mixed disulfide intermediate (MDI). The formation of MDI was monitored with either UV‐Visible spectroscopy by observing an absorbance increase at 412 nm as a chromophore tag, TNB, was released upon reaction between TtSco and TtCuA or by non‐reducing SDS‐PAGE gels. Altering Arg 48, which is adjacent to the attacking cysteine to an Asp has a very large effect on MDI production. TtSco also has metal binding ability, found to bind Cu(I), Cu(II), Ni(II), Cd(II) and Co(II) with varying relative affinities, in addition to disulfide reductase activity. To further characterize MDI formation with TtSco, the impact of conserved residues involved in metal binding on MDI formation were probed and showed that changing the conserved histidine did not affect MDI formation. However, binding of metal to Sco prior to incubating with TtCuA abrogated MDI formation. In characterizing the metal bound protein, preliminary studies to determine the binding affinity of Cu(II) to TtSco have been completed. The impact of conserved cysteines on metal binding has also been studied and the results indicate that one cysteine binds much more tightly to the Cu(II) than the other. While this work allows for a better understanding of TtSco function, much work remains to be done to understand the impacts of metal binding and to further characterize the Sco family of proteins.Support or Funding InformationResearch Corporation (Cottrell College Science Award 7963), the Welch Foundation (W‐0031 [Trinity Chemistry Department]), FASTER grant SURF‐ National Science Foundation DUE S‐STEM Award 1153796 and the Arnold and Mabel Beckman Foundation Beckman Scholars AwardThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.