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Nanoscale Biopolymers with Customizable Properties for Heavy Metal Remediation

Nanoscale Biopolymers with Customizable Properties for Heavy Metal Remediation. U Loi (Ann) Lao, Giri Prabhukumar, Jan Kostal, Mark Matsumoto, Ashok Mulchandani, and Wilfred Chen Chemical and Environmental Engineering University of California, Riverside. Heavy Metal Contamination.

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Nanoscale Biopolymers with Customizable Properties for Heavy Metal Remediation

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  1. Nanoscale Biopolymers with Customizable Properties for Heavy Metal Remediation U Loi (Ann) Lao, Giri Prabhukumar,Jan Kostal, Mark Matsumoto, Ashok Mulchandani, and Wilfred Chen Chemical and Environmental Engineering University of California, Riverside

  2. Heavy Metal Contamination • 2.4 million tons of metal wastes per year from industrial sources • 2 million tons per year from agriculture and domestic waste • Pb2+, Hg2+ and Cd2+ are ranked 2nd, 3rd and 7th, respectively, on the EPA’s priority list

  3. Metal Chelating Polymers • Requires toxic solvents for synthesis • Ultrafiltration is required • Membrane clogging Solution: Develop metal-binding materials that can be recovered by environmental stimuli

  4. Metal Chelating Biopolymers • Based on biological building blocks • Nanoscale biopolymers that are specially pre-programmed within a DNA template • Economically produced by bacteria • Environmentally friendly • Tunable properties based on changes in environmental conditions – pH or temperature

  5. Elastin Biopolymer • Structurally similar to the repeating elastometric peptide sequence of the mammalian protein, elastin • VPGVG are the most frequently repeating units • Undergo a reversible phase transition from water soluble forms into aggregates as the temperature increases High temperature: twisted filament of b-spirals Low temperature: b-turn

  6. Plasmid Enzyme Recombinant plasmid Synthetic gene ELP biopolymers Genetic and Protein Engineering Methodology

  7. Fine tune affinity with different binding sequences Fine tune the transition temperature by controlling amino acid sequence and number of repeating unit (VPGXG)n Customizable Metal Binding Biopolymers Metal Binding Domain Elastin Domain

  8. Cd2+ Cd2+ Cd2+ Cd2+ Cd2+ Cd2+ Regeneration + Cd DT soluble protein insoluble protein Heavy Metal Removal by Tunable Biopolymer

  9. Biopolymer A. ELP38H6 B. ELP58H6 C. ELP78H6 D. ELP78 E. ELP78H12 Elastin Biopolymers with Metal-Binding Affinity A B C D E Kostal et al. Macromolecules, 34, 2257-2261, 2001.

  10. High Temp Low Temp Phase Transition Transition temperatures can be fine tuned from 20 to 40oC by controlling the chain length and salt concentration

  11. Cadmium Removal 50 nmol of biopolymer were incubated with 100 nmol Cd2+

  12. Repeated Cycles of Metal Binding 5 mg added each cycle Cycle 1 Cycle 2 Cycle 3 Cycle 4

  13. Practical Applications: Soil Flushing/Washing • Environmental friendly extractant • Simple separation and recycling In-situ Soil Flushing Ex-situ Soil Washing

  14. Soil-Biopolymer Adsorption Characteristics • Soil-Biopolymer Sorption Characteristics • Freundlich isotherm Log x/m = Log Kf + 1/n Log Ceq Maximum 19% adsorption Biosurfactant – typically in the 70% range Prabhukumar et al. ES&T, 38, 3148-3152, 2004.

  15. 3. Single batch washing with 1.25 mg/ml (0.036 mM) ELP without the histidine tag 1. Single Batch washing with distilled water 2. Two sequential batch washings with distilled water 4. Single batch washing with 0.036 mM EDTA 5. Single batch washing with 1.25 mg/ml ELPH12 8. Two sequential batch washings with 1.25 mg/ml ELPH12 6. Single batch washing with 2.5 mg/ml ELPH12 7. Single batch washing with 5 mg/ml ELPH12 Biopolymer-Cadmium Extraction Studies Less than 10% adsorption

  16. First generation biopolymers • His6 or His12 tag can serve as a simple metal binding domain • Both Tt and metal binding capacity can be easily regulated • Biopolymers can be recycled • Lack specificity and affinity

  17. Customizable Metal Binding Biopolymers Fine tune affinity with different binding sequences Fine tune DT by controlling amino acid sequence and number of repeating unit (VPGXG)n Metal Binding Domain Elastin Domain

  18. Hg2+ Hg2+ Hg0 MerD MerR MerR Bacterial mer Systems OM IM mer genes on E. coli plasmids or transposons MerA MerA MerT MerP MerB organomercury

  19. Hg2+ MerR can serve as a specific mercury binding domain C123 C79 C114 C114 C79 C123 MerR-Hg complex

  20. A B C kDa Production and purification of ELP153-MerR A, cell free extract B, purified protein (80.1 kDa) C, size marker

  21. Binding of Mercury to ELP153-MerR Similar binding from pH 4 to 9

  22. Selective Binding of Mercury by Ela153-MerR Biopolymer Acidic waste water (pH 4) Kostal et al. ES&T, 37, 4457-4462, 2003.

  23. Other Metal-binding Domains Metallothioneins (MTs) MDRNCSCAACDSCTCAGSCKCKECKCTSCKKSCCSCCPVGCAKCAQGCICKGASDKCSCCA Synthetic Phytochelatins (ECs) gECECECECECECECECECECECECECECECECECECECECG Higher affinity toward Hg and Cd

  24. Prefer Hg over Cd Cadmium Binding

  25. Hg2+ Hg2+ Cd2+ EC20 EC20 ELP105K21 ELP105K21 +ΔT1 Cd2+ Histag +ΔT2or NaCl Hg2+ ELP78 Differential Precipitation and Separation – Strategy

  26. Customizable Metal Binding Biopolymers Fine tune affinity with different binding sequences Fine tune DT by controlling amino acid sequence and number of repeating unit (VPGXG)n Metal Binding Domain Elastin Domain

  27. Differential Precipitation and Separation • Differential precipitation and recovery by tuning the elastin composition: Lysine as the guest residues ELP78+ ELP-Lys ELP78 ELP-Lys

  28. ELPKEC20 ELPH12 Differential Precipitation and Separation 1 2 3 4 5 Mixture Heat to 32oC Centrifuge at 32oC Pellet dissolved in cold buffer Supernatant heat to 48oC Centrifuge at 48oC Pellet dissolved in cold buffer • Marker. 4) 48 oC heat pellet • Biopolymer mixture 5) 48 oC supernatant • 3) 32 oC heat spin pellet

  29. Conclusions • Nanoscale ELP biopolymer can be designed with customized properties: • Specificity • Tunable transition • Provide a novel, environmental friendly, and green engineering method for removal of heavy metal from water and soil.

  30. Acknowledgement R829606 Exploratory Research: Nanotechnology

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