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The Science of the Total Environment Aug 2023Anthropogenic activities have increased the dispersal of emerging contaminants (ECs), particularly of parabens, causing an escalation of their presence in wastewater...
Anthropogenic activities have increased the dispersal of emerging contaminants (ECs), particularly of parabens, causing an escalation of their presence in wastewater (WW). Current WW technologies do not present satisfactory efficiency or sustainability in removing these contaminants. However, bioremediation with microalgae-based systems is proving to be a relevant technology for WW polishing, and the use of microalgae-bacteria consortia can improve the efficiency of WW treatment. This work aimed to study dual cultures of selected bacteria (Raoultella ornithinolytica, Acidovorax facilis, Acinetobacter calcoaceticus, Leucobacter sp. or Rhodococcus fascians) and the microalga Chlorella vulgaris in microbial growth and WW bioremediation - removal of methylparaben (MetP) and nutrients. The association with the bacteria was antagonistic for C. vulgaris biomass productivity as a result of the decreased growth kinetics in comparison to the axenic microalga. The presence of MetP did not disturb the growth of C. vulgaris under axenic or co-cultured conditions, except when associated with R. fascians, where growth enhancement was observed. The removal of MetP by the microalga was modest (circa 30 %, with a removal rate of 0.0343 mg/L.d), but increased remarkably when the consortia were used (> 50 %, with an average removal rate > 0.0779 mg/L.d), through biodegradation and photodegradation. For nutrient removal, the consortia were found to be less effective than the axenic microalga, except for nitrogen (N) removal by C. vulgaris w/ R. fascians. The overall results propose that C. vulgaris co-cultivation with bacteria can increase MetP removal, while negatively affecting the microalga growth and the consequent reduction of sludge production, highlighting the potential of microalgae-bacteria consortia for the effective polishing of WW contaminated with parabens.
Topics: Chlorella vulgaris; Wastewater; Coculture Techniques; Parabens; Bacteria; Microalgae
PubMed: 37121314
DOI: 10.1016/j.scitotenv.2023.163746 -
Biotechnology Letters Aug 2021Catalytic promiscuity, or the ability to catalyze a secondary reaction, provides new opportunities for industrial biocatalysis by expanding the range of biocatalytic...
OBJECTIVES
Catalytic promiscuity, or the ability to catalyze a secondary reaction, provides new opportunities for industrial biocatalysis by expanding the range of biocatalytic reactions. Some nitrilases converting nitriles to amides, referred to as the secondary activity, show great potential for amides production. And our goal was exploiting the amide-forming potential of nitrilases.
RESULTS
In this study, we characterized and altered the secondary activity of nitrilase from Acidovorax facilis 72 W (Nit72W) towards different substrates. We increased the secondary activity of Nit72W towards 2-cyanopyridine by 196-fold and created activity toward benzonitrile and p-nitrophenylacetonitrile by modifying the active pocket. Surprisingly, the best mutant, W188M, completely converted 250 mM 2-cyanopyridine to more than 98% 2-picolinamide in 12 h with a specific activity of 90 U/mg and showed potential for industrial applications.
CONCLUSIONS
Nit72W was modified to increase its secondary activity for the amides production, especially 2-picolinamide.
Topics: Aminohydrolases; Bacterial Proteins; Biocatalysis; Comamonadaceae; Picolinic Acids; Protein Engineering; Substrate Specificity
PubMed: 33961157
DOI: 10.1007/s10529-021-03137-7 -
Journal of Microbiology and... May 2019Nitrilase is a valuable type of hydrolase that catalyzes nitriles into carboxylic acid and ammonia. Its applications, however, are severely restricted by the harsh...
Nitrilase is a valuable type of hydrolase that catalyzes nitriles into carboxylic acid and ammonia. Its applications, however, are severely restricted by the harsh conditions of industrial reaction processes. To solve this problem, a nitrilase from 72W was inserted into an shuttle vector for spore surface display. Western blot, enzyme activity measurements and flow cytometric analysis results all indicated a successful spore surface display of the CotB-nit fusion protein. In addition, the optimal catalytic pH value and temperature of the displayed nitrilase were determined to be 7.0 and 50°C, respectively. Moreover, results of reusability tests revealed that 64% of the initial activity of the displayed nitrilase was still retained at the 10 cycle. Furthermore, hydrolysis efficiency of upscale production of cyanocarboxylic acid was significantly higher in the displayed nitrilase-treated group than in the free group expressed by (pET-28a-nit). Generally, the display of 72W nitrilase on the spore surface of may be a useful method for immobilization of enzyme and consequent biocatalytic stabilization.
Topics: Aminohydrolases; Bacillus subtilis; Bacterial Proteins; Comamonadaceae; Enzyme Stability; Enzymes, Immobilized; Escherichia coli; Genetic Vectors; Hydrogen-Ion Concentration; Immobilization; Recombinant Fusion Proteins; Spores, Bacterial; Temperature; Time Factors
PubMed: 30955259
DOI: 10.4014/jmb.1901.01030 -
International Journal of Biological... Jun 2019Nitrilase-catalyzed regioselective hydrolysis of 1-cyanocyclohexaneacetonitrile (1-CHAN) is a green and efficient approach for the preparation of...
Enhanced catalytic stability and reusability of nitrilase encapsulated in ethyleneamine-mediated biosilica for regioselective hydrolysis of 1-cyanocycloalkaneacetonitrile.
Nitrilase-catalyzed regioselective hydrolysis of 1-cyanocyclohexaneacetonitrile (1-CHAN) is a green and efficient approach for the preparation of 1-cyanocyclohexaneacetic acid (1-CHAA), a key precursor for the synthesis of gabapentin. Here, a mesoporous biosilica particles prepared by the ethyleneamine-mediated silicification have been used as carrier for the encapsulation of nitrilase from Acidovorax facilis (NitA). The silica-encapsulated NitA (NitA@silica) with triethylenetetramine as an initiator showed the highest immobilization efficiency (98.3%) and specific activity (672.6 U/g). Both free and encapsulated NitA were optimally active at 40 °C and pH 7.0, however, the encapsulated enzyme exhibited wider optimum temperature range, and enhanced thermal stability compared with the free enzyme. The kinetic parameters K and V for free and encapsulated NitA were calculated to be 141 mM and 9.97 mM min, and 280 mM and 9.02 mM min, respectively. The encapsulated NitA showed good reusability and retained about 94.2% of its initial activity even after 16 cycles of reaction. Also, the storage experiments revealed high activity maintenance of encapsulated NitA after 17-day storage at 4 °C. A preparative scale regioselective hydrolysis of 1-CHAN to 1-CHAA with encapsulated NitA as biocatalyst was carried out in a 2 L stirred bioreactor. The concentration of 1-CHAA reached 152 g/L after 8 h reaction and the conversion was 90.9%. These results showed that the encapsulation of NitA in ethyleneamine-mediated biosilica is an efficient and simple way for preparation of stable nitrilase and have a great potential for application in enzymatic production of carboxylic acids.
Topics: Acetonitriles; Aminohydrolases; Biocatalysis; Capsules; Cyclohexanes; Enzyme Stability; Enzymes, Immobilized; Hydrogen-Ion Concentration; Hydrolysis; Kinetics; Nitriles; Silicon Dioxide; Stereoisomerism; Substrate Specificity; Temperature; Trientine
PubMed: 30807797
DOI: 10.1016/j.ijbiomac.2019.02.131 -
Bioprocess and Biosystems Engineering Mar 2019Nitrilase is the member of carbon-nitrogen hydrogen hydrolase superfamily, which has been widely used for the hydrolysis of nitriles into corresponding carboxylic acids....
Nitrilase is the member of carbon-nitrogen hydrogen hydrolase superfamily, which has been widely used for the hydrolysis of nitriles into corresponding carboxylic acids. But most nitrilases are plagued by product inhibition in the industrial application. In this study, a "super nitrilase mutant" of nitrilase with high activity, thermostability and improved product tolerance from Acidovorax facilis ZJB09122 was characterized. Then, an efficient process was developed by employing the whole cell of recombinant E. coli for the conversion of high concentration of 1-cyanocyclohexylacetonitrile-to-1-cyanocyclohexaneacetic acid, an important intermediate of gabapentin. Under the optimized conditions, the higher substrate concentrations such as 1.3 M, 1.5 M and 1.8 M could be hydrolyzed by 13.58 g DCW/L with outstanding productivity (> 740 g/L/day). This study developed a highly efficient bioprocess for the preparation of 1-cyanocyclohexaneacetic acid which has the great potential for industrial application.
Topics: Aminohydrolases; Bacterial Proteins; Comamonadaceae; Escherichia coli; Mutation; Nitriles; Recombinant Proteins
PubMed: 30488321
DOI: 10.1007/s00449-018-2049-2 -
International Journal of Biological... Sep 2018Iminodiacetic acid (IDA) is widely used as an intermediate in the manufacturing of chelating agents, glyphosate herbicides and surfactants. To improve activity and...
Iminodiacetic acid (IDA) is widely used as an intermediate in the manufacturing of chelating agents, glyphosate herbicides and surfactants. To improve activity and tolerance to the substrate for IDA production, Acidovorax facilis nitrilase was selected for further modification by the gene site saturation mutagenesis method. After screened by a two-step screening method, the best mutant (Mut-F168V/T201N/S192F/M191T/F192S) was selected. Compared to the wild-type nitrilase, Mut-F168V/T201N/S192F/M191T/F192S showed 136% improvement in specific activity. Co stimulated nitrilase activity, whereas Cu, Zn and Tween 80 showed a strong inhibitory effect. The V and k of Mut-F168V/T201N/S192F/M191T/F192S were enhanced 1.23 and 1.23-fold, while the K was decreased 1.53-fold. The yield of Mut-F168V/T201N/S192F/M191T/F192S with 453.2 mM of IDA reached 71.9% in 5 h when 630 mM iminodiacetonitrile was used as substrate. This study indicated that mutant nitrilase obtained in this study is promising in applications for the upscale production of IDAN.
Topics: Amino Acid Substitution; Aminohydrolases; Bacterial Proteins; Comamonadaceae; Mutagenesis, Site-Directed; Recombinant Proteins
PubMed: 29753012
DOI: 10.1016/j.ijbiomac.2018.05.045 -
Journal of Hazardous Materials Apr 2018The contamination of the environment by U may affect plant life and consequently may have an impact on animal and human health. The present work describes U(VI)...
The contamination of the environment by U may affect plant life and consequently may have an impact on animal and human health. The present work describes U(VI) sequestration by Acidovorax facilis using a multidisciplinary approach combining wet chemistry, transmission electron microscopy, and spectroscopy methods (e.g. cryo-time resolved laser-induced fluorescence spectroscopy, extended X-ray absorption fine structure spectroscopy, and in-situ attenuated total reflection Fourier transform infrared spectroscopy). This bacterial strain is widely distributed in nature including U-contaminated sites. In kinetic batch experiments cells of A. facilis were contacted for 5 min to 48 h with 0.1 mM U(VI). The results show that the local coordination of U species associated with the cells depends upon time contact. U is bound mainly to phosphate groups of lipopolysaccharide (LPS) at the outer membrane within the first hour. And, that both, phosphoryl and carboxyl functionality groups of LPS and peptidoglycan of A. facilis cells may effectuate the removal of high U amounts from solution at 24-48 h of incubation. It is clearly demonstrated that A. facilis may play an important role in predicting the transport behaviour of U in the environment and that the results will contribute to the improvement of bioremediation methods of U-contaminated sites.
Topics: Biodegradation, Environmental; Comamonadaceae; Lipopolysaccharides; Radioactive Pollutants; Uranium; Water Purification
PubMed: 29324323
DOI: 10.1016/j.jhazmat.2017.12.030 -
Scientific Reports Sep 2017Bacteria are essential in arsenic cycling. However, few studies have addressed 16S rRNA and arsenic-related functional gene diversity in long-term arsenic-contaminated...
Bacteria are essential in arsenic cycling. However, few studies have addressed 16S rRNA and arsenic-related functional gene diversity in long-term arsenic-contaminated tropical sediment. Here, using culture-based, metagenomic and computational approaches, we describe the diversity of bacteria, genes and enzymes involved in AsIII and AsV transformation in freshwater sediment and in anaerobic AsIII- and AsV-enrichment cultures (ECs). The taxonomic profile reveals significant differences among the communities. Arcobacter, Dechloromonas, Sedimentibacter and Clostridium thermopalmarium were exclusively found in ECs, whereas Anaerobacillus was restricted to AsV-EC. Novel taxa that are both AsV-reducers and AsIII-oxidizers were identified: Dechloromonas, Acidovorax facilis, A. delafieldii, Aquabacterium, Shewanella, C. thermopalmarium and Macellibacteroides fermentans. Phylogenic discrepancies were revealed among the aioA, arsC and arrA genes and those of other species, indicating horizontal gene transfer. ArsC and AioA have sets of amino acids that can be used to assess their functional and structural integrity and familial subgroups. The positions required for AsV reduction are conserved, suggesting strong selective pressure for maintaining the functionality of ArsC. Altogether, these findings highlight the role of freshwater sediment bacteria in arsenic mobility, and the untapped diversity of dissimilatory arsenate-reducing and arsenate-resistant bacteria, which might contribute to arsenic toxicity in aquatic environments.
Topics: Anaerobiosis; Arsenic; Bacteria; Biotransformation; Cluster Analysis; DNA, Bacterial; DNA, Ribosomal; Enzymes; Fresh Water; Genetic Variation; Geologic Sediments; Metabolic Networks and Pathways; Phylogeny; RNA, Ribosomal, 16S; Sequence Analysis, DNA; Water Pollutants, Chemical
PubMed: 28894204
DOI: 10.1038/s41598-017-11548-8 -
Journal of Hazardous Materials Nov 2016The former uranium mine Königstein (Saxony, Germany) is currently in the process of remediation by means of controlled underground flooding. Nevertheless, the flooding...
The former uranium mine Königstein (Saxony, Germany) is currently in the process of remediation by means of controlled underground flooding. Nevertheless, the flooding water has to be cleaned up by a conventional wastewater treatment plant. In this study, the uranium(VI) removal and tolerance mechanisms of the gram-negative betaproteobacterium Acidovorax facilis were investigated by a multidisciplinary approach combining wet chemistry, flow cytometry, and microscopy. The kinetics of uranium removal and the corresponding mechanisms were investigated. The results showed a biphasic process of uranium removal characterized by a first phase where 95% of uranium was removed within the first 8h followed by a second phase that reached equilibrium after 24h. The bacterial cells displayed a total uranium removal capacity of 130mgU/g dry biomass. The removal of uranium was also temperature-dependent, indicating that metabolic activity heavily influenced bacterial interactions with uranium. TEM analyses showed biosorption on the cell surface and intracellular accumulation of uranium. Uranium tolerance tests showed that A. facilis was able to withstand concentrations up to 0.1mM. This work demonstrates that A. facilis is a suitable candidate for in situ bioremediation of flooding water in Königstein as well as for other contaminated waste waters.
Topics: Adsorption; Biodegradation, Environmental; Biomass; Comamonadaceae; Flow Cytometry; Germany; Microbial Sensitivity Tests; Microbial Viability; Microscopy, Electron, Transmission; Microscopy, Fluorescence; Mining; Uranium Compounds; Wastewater; Water Pollutants, Radioactive
PubMed: 27262280
DOI: 10.1016/j.jhazmat.2016.05.062 -
Otology & Neurotology : Official... Sep 2013To describe the microbial flora associated with cochlear implants (CIs) removed for infectious and noninfectious indications.
OBJECTIVE
To describe the microbial flora associated with cochlear implants (CIs) removed for infectious and noninfectious indications.
STUDY DESIGN
Prospective, controlled.
SETTING
Academic, tertiary medical center.
PATIENTS
All patients undergoing CI removal.
INTERVENTION
CIs were removed with aseptic technique and processed for microbial identification.
MAIN OUTCOME MEASURE
CI microbes were identified with routine culture and speciation and 16s deoxyribonucleic acid 454-pyrosequencing.
RESULTS
All CIs had evidence of microbes. Propionibacterium acnes and Acidovorax facilis were more common on noninfected CIs (p = 0.005, 0.031). Staphylococcus aureus was more common on infected CIs (p = 0.003). The microbial profiles associated with CI infection were different from, but overlapped with those of noninfected CIs. Microbial culture with selective media identified pathogens not identified on pyrosequencing.
CONCLUSION
Bacteria are present on all CIs, both with and without evidence of clinical infection, but species differ with clinical status. Empiric therapy for CI infections should include coverage for S. aureus. Gene pyrosequencing and selective culture techniques may yield complementary results that may impact the management of CI infections.
Topics: Adolescent; Adult; Aged; Aged, 80 and over; Bacterial Infections; Biofilms; Child; Child, Preschool; Cochlear Implants; DNA, Bacterial; Data Interpretation, Statistical; Device Removal; Ear, Inner; Female; Genes, Bacterial; Humans; Magnets; Male; Microscopy, Electron, Scanning; Middle Aged; Polymerase Chain Reaction; Propionibacterium acnes; Prosthesis-Related Infections; Risk Factors; Sequence Analysis, DNA; Staphylococcus aureus; Young Adult
PubMed: 23921938
DOI: 10.1097/MAO.0b013e3182941101