Gold Biomineralization Research Articles

Functional and physicochemical characterization of marine exopolysaccharide derived from the novel bacterium Algoriphagus sp. K5R and its application towards biomineralization of gold and silver

AbstractThe widespread use of synthetic polymers in various industries has raised worldwide concerns regarding their ecological impact and effects on human health. As a result, biopolymers have emerged as a promising alternative. Among them, exopolysaccharides (EPSs) produced by microbes from terrestrial niches have been extensively studied. However, recent reports have indicated that microbes from marine environments can also produce unique EPSs that could serve as sustainable substitutes to meet the escalating demand for biopolymers. The present study, for the first time, reports EPS production from novel marine bacterium Algoriphagus sp. K5R for sustainable application development. Interestingly, physicochemical analyses suggest that EPS K5R is a high molecular weight (1190.63 kDa) heteropolysaccharide composed of galacturonic acid, glucose and mannose. To evaluate EPS production, growth and fermentation kinetics were performed, which revealed that it was a primary metabolite having a maximum production of 4.79 ± 0.57 g L−1 with 2% (w/v) glucose. Moreover, EPS K5R exhibits exceptional functional properties, namely high water‐holding capacity (720% ± 80.29%) and oil‐holding capacity (493.33% ± 49.74% for coconut oil and 533.32% ± 17.47% for olive oil), and non‐Newtonian pseudo‐plastic behavior which render it a promising candidate for application in the cosmetics and food industries. In fact, its anti‐oxidant capabilities make it an ideal biological reducing agent for metal nanoparticle synthesis. Overall, this study highlights the potential of marine EPSs for a diverse array of industrial applications. © 2024 Society of Chemical Industry.

Synthetic Biology Toolbox, Including a Single-Plasmid CRISPR-Cas9 System to Biologically Engineer the Electrogenic, Metal-Resistant Bacterium Cupriavidus metallidurans CH34.

Cupriavidus metallidurans CH34 exhibits extraordinary metabolic versatility, including chemolithoautotrophic growth; degradation of BTEX (benzene, toluene, ethylbenzene, xylene); high resistance to numerous metals; biomineralization of gold, platinum, silver, and uranium; and accumulation of polyhydroxybutyrate (PHB). These qualities make it a valuable host for biotechnological applications such as bioremediation, bioprocessing, and the generation of bioelectricity in microbial fuel cells (MFCs). However, the lack of genetic tools for strain development and studying its fundamental physiology represents a bottleneck to boosting its commercial applications. In this study, inducible and constitutive promoter libraries were built and characterized, providing the first comprehensive list of biological parts that can be used to regulate protein expression and optimize the CRISPR-Cas9 genome editing tools for this host. A single-plasmid CRISPR-Cas9 system that can be delivered by both conjugation and electroporation was developed, and its efficiency was demonstrated by successfully targeting the pyrE locus. The CRISPR-Cas9 system was next used to target candidate genes encoding type IV pili, hypothesized by us to be involved in extracellular electron transfer (EET) in this organism. Single and double deletion strains (ΔpilA, ΔpilE, and ΔpilAE) were successfully generated. Additionally, the CRISPR-Cas9 tool was validated for constructing genomic insertions (ΔpilAE::gfp and ΔpilAE::λPrgfp). Finally, as type IV pili are believed to play an important role in extracellular electron transfer to solid surfaces, C. metallidurans CH34 ΔpilAE was further studied by means of cyclic voltammetry using disposable screen-printed carbon electrodes. Under these conditions, we demonstrated that C. metallidurans CH34 could generate extracellular currents; however, no difference in the intensity of the current peaks was found in the ΔpilAE double deletion strain when compared to the wild type. This finding suggests that the deleted type IV pili candidate genes are not involved in extracellular electron transfer under these conditions. Nevertheless, these experiments revealed the presence of different redox centers likely to be involved in both mediated electron transfer (MET) and direct electron transfer (DET), the first interpretation of extracellular electron transfer mechanisms in C. metallidurans CH34.

Supergene gold mineralization from exploited placer deposits at Dziwiszów in the Sudetes (NE Bohemian Massif, SW Poland)

Representative ore samples collected from outcrops in the exploited Dziwiszów placer gold deposit in Lower Silesia offer the opportunity to study supergene transformations which probably occurred over historic periods. The ore samples are alloys of gold and silver, with trace amounts of Hg and Pd. There are three distinct sub-types of detrital gold: (1) flaky, (2) nugget-like; and (3) irregular/craggy grains, respectively. All samples record the features of supergene processes such as dissolution textures, as well as the formation and aggregation of authigenic nano- and microparticles of native gold. Surface authigenic morphotypes include nano-particles of semi-spherical or blebby native gold, as well as overgrowths of crystalline phases of platy gold and micro-aggregates of bacteriomorphic Au, which provide evidence for microbial gold biomineralization. The growth history of individual native gold particles is documented by nano- to micro-textural investigations of sample grains ranging from isolated semi-spherical nanoparticles through agglomerates to irregularly shaped platy micro-nuggets of gold. The presence of nano- to microphase native gold embedded in fine-grained assemblages of clayish masses and the absence of any evidence for abrasion of individual grains suggest the authigenic origin of blebby native gold particles and overgrowths of platy native gold grains. Hence, we propose that these particles were formed in situ. Chemical transfer may be related to both the precipitation of Au from a colloidal solution, as well as to its adsorption by clay minerals and surface precipitation on particles of the mineral substrate.

Intratumoral generation of photothermal gold nanoparticles through a vectorized biomineralization of ionic gold

Various cancer cells have been demonstrated to have the capacity to form plasmonic gold nanoparticles when chloroauric acid is introduced to their cellular microenvironment. But their biomedical applications are limited, particularly considering the millimolar concentrations and longer incubation period of ionic gold. Here, we describe a simplistic method of intracellular biomineralization to produce plasmonic gold nanoparticles at micromolar concentrations within 30 min of application utilizing polyethylene glycol as delivery vector for ionic gold. We have characterized this process for intracellular gold nanoparticle formation, which progressively accumulates proteins as the ionic gold clusters migrate to the nucleus. This nano-vectorized application of ionic gold emphasizes its potential biomedical opportunities while reducing the quantity of ionic gold and required incubation time. To demonstrate its biomedical potential, we further induce in-situ biosynthesis of gold nanoparticles within MCF7 tumor mouse xenografts which is followed by its photothermal remediation.

Detecting Gold Biomineralization by Delftia acidovorans Biofilms on a Quartz Crystal Microbalance.

The extensive use of gold in sensing, diagnostics, and electronics has led to major concerns in solid waste management since gold and other heavy metals are nonbiodegradable and can easily accumulate in the environment. Moreover, gold ions are extremely reactive and potentially harmful for humans. Thus, there is an urgent need to develop reliable methodologies to detect and possibly neutralize ionic gold in aqueous solutions and industrial wastes. In this work, by using complementary measurement techniques such as quartz crystal microbalance (QCM), atomic force microscopy, crystal violet staining, and optical microscopy, we investigate a promising biologically induced gold biomineralization process accomplished by biofilms of bacterium Delftia acidovorans. When stressed by Au3+ ions, D. acidovorans is able to neutralize toxic soluble gold by excreting a nonribosomal peptide, which forms extracellular gold nanonuggets via complexation with metal ions. Specifically, QCM, a surface-sensitive transducer, is employed to quantify the production of these gold complexes directly on the D. acidovorans biofilm in real time. Detailed kinetics obtained by QCM captures the condition for maximized biomineralization yield and offers new insights underlying the biomineralization process. To the best of our knowledge, this is the first study providing an extensive characterization of the gold biomineralization process by a model bacterial biofilm. We also demonstrate QCM as a cheap, user-friendly sensing platform and alternative to standard analytical techniques for studies requiring high-resolution quantitative details, which offers promising opportunities in heavy-metal sensing, gold recovery, and industrial waste treatment.

Gold Biomineralization on Bacterial Biofilms for Leaching of Au3+ Damages Eukaryotic Cells.

Heavy metals not only pollute the environment but also are health and environmental hazard. Bacteria constitute inexpensive and eco-friendly material to eliminate and recycle heavy metals via biomineralization and biosorption. However, the effect of metal biomineralization in bacterial biofilms on the ecological balance of bacteria and infectious diseases is unclear. This study aimed to explore the interaction between a eukaryotic cell line HEK293T and mineralized Escherichia coli, using a model of gold biomineralization on E. coli biofilms (E. coli-Au). In our present model, bacterial activity was not disrupted and bacterial adhesion and invasion were enhanced. E. coli-Au invaded the cytoplasm and nuclei of HEK293T cells and damaged them via intracellular growth and multiplication. The present findings indicate that metal biomineralization in bacterial biofilms for leaching of heavy metal ions is hazardous to eukaryotic cells and even human health.

Selective Mineralization and Recovery of Au(III) from Multi-Ionic Aqueous Systems by Bacillus licheniformis FZUL-63

The recovery of precious metals is a project with both economic and environmental significance. In this paper, how to use bacterial mineralization to selectively recover gold from multi-ionic aqueous systems is presented. The Bacillus licheniformis FZUL-63, isolated from a landscape lake in Fuzhou University, was shown to selectively mineralize and precipitate gold from coexisting ions in aqueous solution. The removal of Au(III) almost happened in the first hour. Scanning electron microscope with X-ray energy dispersive spectroscopy (SEM/EDS-mapping) results and fourier transform infrared spectroscopy (FTIR) data show that the amino, carboxyl, and phosphate groups on the surface of the bacteria are related to the adsorption of gold ions. X-ray photoelectron spectroscopy (XPS) results implied that Au(III) ions were reduced to those that were monovalent, and the Au(I) was then adsorbed on the bacterial surface at the beginning stage (in the first hour). X-ray diffraction (XRD) results showed that the gold biomineralization began about 10 h after the interaction between Au(III) ions and bacteria. Au(III) mineralization has rarely been influenced by other co-existing metal ions. Transmission electron microscope (TEM) analysis shows that the gold nanoparticles have a polyhedral structure with a particle size of ~20 nm. The Bacillus licheniformis FZUL-63 could selectively mineralize and recover 478 mg/g (dry biomass) gold from aqua regia-based metal wastewater through four cycles. This could be of great potential in practical applications.

Extracellular Biomineralization of Gold by Delftia tsuruhatensis GX-3 Isolated from a Heavy Metal Contaminated Paddy Soil

The use of microorganisms in geochemical gold recycling was considered recently. To explore the feasibility of gold biomineralization in this study, bacteria GX-3 were isolated from a heavy metal contaminated paddy soil in Guangxi province, China. This sulfur-reducing bacteria, Delftia tsuruhatensis, can survive in toxic Au3+ solution. Alkaline conditions (pH 8.0) and 35 °C were optimal for GX-3 cultivation. Nanoparticle formation was promoted by the coexistence of Fe3+. Field-emission scanning electron microscopy and transmission electron microscopy (TEM) images of GX-3 showed the presence of spherical particles, ranging from 50 to 100 and 200 to 500 nm in diameter, respectively, that stuck together and distributed around the surface of the bacteria as well as outside the cell. These nanoparticles were identified as gold according to the TEM–energy dispersive X-ray spectroscopy analysis. Here, bacterial extracellular solute-binding protein and/or porin should account for the gold nanoparticle formation and the detoxification in aqueous system as it transports electrons to Au3+ for nanoparticle precipitation. The ubiquitous nature of bacteria in near-surface environments and the rapid precipitation of gold by the sulfur-reducing bacterium suggest that these bacteria could be responsible for the wide bioaccumulation of gold in natural systems.

Nanoforms of gold from abandoned placer deposits of Wądroże Wielkie, Lower Silesia, Poland – The evidence of authigenic gold mineralization

Being the most recently exposed, gold grains from historical tailings of underground and surface workings offer the opportunity to study supergene transformations occurring over historic periods. In this study, gold grains were collected from the abandoned, historically mined deposits of Wądroże Wielkie at Lower Silesia.All three distinct gold sub-types of detrital gold display surface morphotypes and internal textures of Au and Ag dissolution indicative of supergene gold modification, as well as authigenic Au nanoparticle formation and aggregation. The indicative morphotypes include nano-particulate semispherical or bubble-like gold and aggregates of bacteriomorphic gold that could be evidence for microbial gold biomineralization. SEM investigation of the microtopography and internal texture of gold grains revealed indications of post-depositional dissolution of Au-Ag alloys and redistribution of these components within the sediment. Some of seed and bubble-like growths are also embedded in clayey (kaolinite) masses within crevices suggesting that dissolution/re-precipitation processes occur at the gold grain interface.Authigenic gold occurs as overgrowths and aggregates of nearly pure gold on a detrital Au-Ag alloy that are readily removed during transport, but are replaced by re-precipitation occurring over historic rather than geological periods.

Biominerized gold-Hemin@MOF composites with peroxidase-like and gold catalysis activities: A high-throughput colorimetric immunoassay for alpha-fetoprotein in blood by ELISA and gold-catalytic silver staining

Catalytic hemin was first remolded with biomineralized gold to form Hemin-Au core and further encapsulated into the Tb metal organic framework (MOF) matrix. The so obtained Hemin-Au@MOF composites could possess high environmental stability and especially double catalysis activities of enhanced peroxidase-like and gold catalysis. Steady state kinetics studies indicate that Hemin-Au@MOF could present much higher intrinsic catalysis and substrate affinity (lower Michaelis constants) than native hemin and even natural peroxidase. Furthermore, the catalytic composites were employed for labeling antibodies towards the sandwiched immunoassays for alpha-fetoprotein (AFP) as a model biomarker of cancers. The results indicate that the Hemin-Au@MOF-based immunoassays could be conducted by two signal amplification ways of hemin-catalyzed chromogenic reaction and gold-catalyzed silver staining, enabling the detection of AFP in blood with the level down to 0.020 ng mL−1. Importantly, the proposed fabrication protocol combining the gold biomineralization and MOF encapsulation routes may be extended for the fabrication of a variety of enzymes or enzymatic mimics with the improved environmental stability and multiple catalysis activities for the wide catalysis applications.

Extraction of gold (Au) particles from sea water by Delftia Acidovorans microbes

Gold-mining activities have been an issue, especially when it involves in contamination of chemicals, for example arsenic and mercury. However, despite of these hazards, gold-mining activities are still being conducted. This is because the gold is worth, regardless of the problems. Gold-mining, as known needs a very large area of land, or site plant. Vast amount of labor force, mechanical force and fund are a must in order for the mining process to be continued. High demand of gold, made gold-mining industries as ones of the most profitable industries in the world. Thus, this has encouraged another alternative way to extract gold. At the mining site, researchers found that biomineralization of gold by Delftia acidovorans can be conducted. How it is done still cannot be understood. It is said that the bacteria secretes secondary metabolites, Delftibactin as a defensive mechanism against the toxicity of the soluble gold. Researchers try to find another source of elemental gold besides of the earth’s core. The options are either lava of a volcano or ocean. Here, the focus is seawater. The problem of seawater is that its composition still not yet to be proved. Dissolve gold existed as gold chloride in seawater, but in a very small amount. So, the gold separation should be focused, in order to make this process to be a successful one. Factors such as depth, climate, region, temperature need to be considered. If this difference affecting the separating process, standardized seawater composition have to be proposed.

Biomineralization of gold by Mucor plumbeus: The progress in understanding the mechanism of nanoparticles' formation.

This contribution describes the deposition of gold nanoparticles by microbial reduction of Au(III) ions using the mycelium of Mucor plumbeus. Biosorption as the major mechanism of Au(III) ions binding by the fungal cells and the reduction of them to the form of Au(0) on/in the cell wall, followed by the transportation of the synthesized gold nanoparticles to the cytoplasm, is postulated. The probable mechanism behind the reduction of Au(III) ions is discussed, leading to the conclusion that this process is nonenzymatic one. Chitosan of the fungal cell wall is most likely to be the major molecule involved in biomineralization of gold by the mycelium of M. plumbeus. Separation of gold nanoparticles from the cells has been carried out by the ultrasonic disintegration and the obtained nanostructures were characterized by UV-vis spectroscopy and transmission electron micrograph analysis. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1381-1392, 2017.

Reconstituting redox active centers of heme-containing proteins with biomineralized gold toward peroxidase mimics with strong intrinsic catalysis and electrocatalysis for H2O2 detection

A facile and efficient enzymatic reconstitution methodology has been proposed for high-catalysis peroxidase mimics by remolding the redox active centers of heme-containing proteins with the in-site biomineralized gold using hemoglobin (Hb) as a model. Catalytic hemin (Hem) was extracted from the active centers of Hb for the gold biomineralization and then reconstituted into apoHb to yield the Hem-Au@apoHb nanocomposites showing dramatically improved intrinsic catalysis and electrocatalysis over natural Hb and Hem. The biomineralized gold, on the one hand, would act as “nanowires” to promote the electron transferring of the nanocomposites. On the other hand, it would create a reactivity pathway to pre-organize and accumulate more substrates towards the active sites of the peroxidase mimics. Steady-state kinetics studies indicate that Hem-Au@apoHb could present much higher substrate affinity (lower Michaelis constants) and intrinsic catalysis even than some natural peroxidases. Moreover, the application feasibility of the prepared artificial enzymes was demonstrated by colorimetric assays and direct electrocatalysis for H2O2 sensing, showing a detection limitation low as 0.45μM. Importantly, such a catalysis active-center reconstitution protocol may circumvent the substantial improvement of the intrinsic catalysis and electrocatalysis of diverse heme-containing proteins or enyzmes toward the extensive applications in the chemical, enviromental, and biomedical catalysis fields.

Multi-analytical approach to understand biomineralization of gold using rice bran: A novel and economical route

Biomimetic fabrication of gold nanoparticles (AuNP) has received significant attention worldwide. However, most of the reports were unable to clearly indicate the responsible biomolecule involved in the production of AuNP. The present study aimed to address this challenge by critically identifying the active component of rice bran responsible for the biomineralization of gold. Fractionation of rice bran extract by HPLC and further characterization using LC-MS revealed that ferulic acid is the functional molecule responsible for the bioconversion of Au3+ to Au0. Real time monitoring of heat changes by isothermal titration calorimetry indicated biosynthesis of AuNP was exothermic in nature. Additionally, reaction between ferulic acid and HAuCl4 for AuNP formation was observed to follow first order kinetics and finally various thermodynamic parameters were estimated by measuring residual ferulic acid concentration. Combinatorial observation of all these studies might lead the way to decipher the exact mechanism of nanoparticle biosynthesis.

Gold nanoparticle formation via microbial metallophore chemistries

Microbes are evolved to live in almost every environment and often produce small molecules to protect themselves, communicate, and sequester nutrients and minerals from their surroundings. Our recent discovery that the gold-associated microbe, Delftia acidovorans, overcomes Au3+ toxicity through complexation and precipitation of soluble gold by the nonribosomal peptide metallophore, delftibactin, revealed a new mechanism for gold biomineralization and a possible industrial application for gold sequestration. In this work, we have sought to define more the scope of nanoparticle formation by other microbial metallophores and the physical properties of gold nanoparticles created by microbial natural products (hydroxamates, catechols, citrates, mixed ligands) and synthetics.

Facile and rapid thermo-regulated biomineralization of gold by pullulan and study of its thermodynamic parameters

A novel method for the production of gold nanoparticles (AuNPs) using pullulan as reducing and stabilizing agent has been developed. Quasi-spherical shaped AuNPs in the range of 50–100nm were produced at three different temperature regimes 80°C, 90°C and 100°C as characterized using UV–vis spectrophotometer, TEM and DLS. Study of reaction kinetics and thermodynamic parameters indicated that the reaction between pullulan and chloroauric acid for AuNPs formation followed first order reaction kinetics and higher temperature was favorable for the synthesis of smaller sized AuNPs. FT-IR data analyses, provided an insight towards the mechanism of gold nanoparticle formation which suggested that, the free CH2OH groups of pullulan molecule were oxidized to carboxylate ions resulted in formation of AuNPs whereas the basic skeletal structure of pullulan remained unaltered. This study may open up new avenues for synthesis of tailor made biogenic AuNPs with possible application in biomedical field.

Geobiological Cycling of Gold: From Fundamental Process Understanding to Exploration Solutions

Microbial communities mediating gold cycling occur on gold grains from (sub)-tropical, (semi)-arid, temperate and subarctic environments. The majority of identified species comprising these biofilms are β-Proteobacteria. Some bacteria, e.g., Cupriavidus metallidurans, Delftia acidovorans and Salmonella typhimurium, have developed biochemical responses to deal with highly toxic gold complexes. These include gold specific sensing and efflux, co-utilization of resistance mechanisms for other metals, and excretion of gold-complex-reducing siderophores that ultimately catalyze the biomineralization of nano-particulate, spheroidal and/or bacteriomorphic gold. In turn, the toxicity of gold complexes fosters the development of specialized biofilms on gold grains, and hence the cycling of gold in surface environments. This was not reported on isoferroplatinum grains under most near-surface environments, due to the lower toxicity of mobile platinum complexes. The discovery of gold-specific microbial responses can now drive the development of geobiological exploration tools, e.g., gold bioindicators and biosensors. Bioindicators employ genetic markers from soils and groundwaters to provide information about gold mineralization processes, while biosensors will allow in-field analyses of gold concentrations in complex sampling media.

Biosynthesis of gold and silver nanoparticles using a novel marine strain of Stenotrophomonas

The present study aims at exploiting marine microbial diversity for biosynthesis of metal nanoparticles and also investigates role of microbial proteins in the process of bio-mineralization of gold and silver. This is the first report for concurrent production of gold and silver nanoparticles (AuNPs and AgNPs) by extracellular secretion of a novel strain of Stenotrophomonas, isolated from Indian marine origin. This novel strain has faster rate kinetics for AgNPs synthesis than any other organism reported earlier. The nanoparticles were further characterized using UV-vis spectrophotometer, TEM, DLS and EDAX confirming their size ranging from 10-50 nm and 40-60 nm in dimensions for AuNPs and AgNPs, respectively. TEM analysis indicated formation of multi-shaped nanoparticles with heterogeneous size distribution in both the cases. Finally, the SDS-PAGE analysis of extracellular media supernatant suggested a potential involvement of certain low molecular weight secretory proteins in AuNPs and AgNPs biosynthesis.

Concurrent zero-dimensional and one-dimensional biomineralization of gold from a solution of Au3+ and bovine serum albumin

A technique was developed for preparing a novel material that consists of gold nanoparticles trapped within a fiber of unfolded proteins. These fibers are made in an aqueous solution that contains HAuCl4 and the protein, bovine serum albumin (BSA). By changing the ratio of gold to BSA in solution, two different types of outcomes are observed. At lower gold to BSA ratios (30–120), a purple solution results after heating the mixture at 80 °C for 4 h. At higher gold to BSA ratios (130–170), a clear solution containing purple fibers results after heating the mixture at 80 °C for 4 h. UV–Vis spectroscopy and light scattering techniques show growth in nanocolloid size as gold to BSA ratio rises above 100. Data indicate that, for the higher gold to BSA ratios, the gold is sequestered within the solid material. The material mass, visible by eye, appears to be an aggregation of smaller individual fibers. Scanning electron microscopy and transmission electron microscopy indicate that these fibers are primarily one-dimensional aggregates, which can display some branching, and can be as narrow as 400 nm in size. The likely mechanism for the synthesis of the novel material is discussed.

Gold biomineralization by a metallophore from a gold-associated microbe

Microorganisms produce and secrete secondary metabolites to assist in their survival. We report that the gold resident bacterium Delftia acidovorans produces a secondary metabolite that protects from soluble gold through the generation of solid gold forms. This finding is the first demonstration that a secreted metabolite can protect against toxic gold and cause gold biomineralization.