Biomining

'''A Separation Problem''': Our access to easily extractable copper is gradually diminishing as demands for copper continues to grow worldwide. To meet these demands, non-traditional, metallurgically challenging deposits are expected to become more prevalent, thereby forcing us to deal with more complex, and lower-grade ores containing higher levels of impurities. Arsenic-challenged deposits is a concern for the copper mining industry as arsenic produces hazardous fumes and oxide dusts during the smelting process. Smelting with arsenic therefore poses a significant risk to the health of the workers and to the environment. Furthermore, safe removal and disposal of stabilized arsenic is often difficult and costly. With stricter legislation in place, the mining industry is facing increasing pressure to progressively reduce the amount of allowable arsenic concentrations for smelters. Consequently, fines and penalties are set for arsenic concentrations exceeding 0.2%, while ores past 0.5% arsenic concentrations are rejected by smelters. Therefore, there is increasing precedence for removing arsenic-containing minerals such as enargite (Cu3AsS4)from relevant sulfide minerals, such as chalcopyrite (CuFeS2), during the flotation process. However, separation is often conflicted with shared flotation properties between enargite and the associated valuable minerals. The mining industry has developed several methods for dealing with arsenic impurities, which includes precipitation with scorodite and pressure hydrometallurgical procedures (150C and 1380kPa) for processing high concentration of arsenic while extracting copper in parallel. However, many procedures requires an enormous amount of energy or additional acidic chemicals to help selectively separate arensic-containing minerals in the ore mixtures.

'''Our solution''': We, the UBC iGEM team, feel that separation and enrichment can be done in other ways that does not rely on energy or chemically-intensive methods. Our synbio solution involves the use of small surface heptapeptides and octapeptides, which have previously been demonstrated in M13 bacteriophage by Curtis et al. to selectively bind to chalcopyrite(3). Three peptides, labelled WSD-1 (TPTTYKV), WSD-2 (DSQKTNPS), and WSD-3 (DPIKHTSG), have been identified for binding. However, operating with bacteriophage is not feasible for large scale operations in mining as it is difficult to scale up titers to compensate for the smaller surface area available for binding in bacteriophage. Our idea is to operate these peptides in bacteria which have a larger surface area and are much more responsive to stimuli. These peptides must be accessible on the surface. Therefore, we have chosen ''Caulobacter crescentus'' as the chosen chassis given that it contains a S-protein layer in which we can express our peptides. A developed kit for protein secretion and display of peptides on the cell surface S-layer in ''Caulobacter crescentus'' (via cloning to the S-layer gene sequence) can be found at [http://ubc.flintbox.com/public/project/1487/ Caulobacter S-layer Kits] A novel protein secretion and display system, using Caulobacter crescentus, that provides scalable, inexpensive secretion of recombinant proteins (via fusion with a C-terminal secretion signal) or display of peptides or proteins on the cell surface (via insertion into the complete S-layer gene sequence). Caulobacter expressing the immunoglobulin G-binding domain of Streptococcal Protein G is available for purchase. The end goals of this project are to bind selectively to chalcopyrite from ore slurries containing chalcopyrite and impurities such as enargite. goal is to sink these Here we show the first application of phage biotechnology to the processing of economically important minerals in ore slurries. A random heptapeptide library was screened for peptide sequences that bind selectively to the minerals sphalerite (ZnS) and chalcopyrite (CuFeS2). After several rounds of enrichment, cloned phage containing the surface peptide loops KPLLMGS and QPKGPKQ bound specifically to sphalerite. Phage containing the peptide loop TPTTYKV bound to both sphalerite and chalcopyrite. By using an enzyme-linked immunosorbant assay (ELISA), the phage was characterized as strong binders compared to wild-type phage. Specificity of binding was confirmed by immunochemical visualization of phage bound to mineral particles but not to silica (a waste mineral) or pyrite. The current study focused primarily on the isolation of ZnS-specific phage that could be utilized in the separation of sphalerite from silica. At mining sites where sphalerite and chalcopyrite are not found together in natural ores, the separation of sphalerite from silica would be an appropriate enrichment step. At mining sites where sphalerite and chalcopyrite do occur together, more specific phage would be required. This bacteriophage has the potential to be used in a more selective method of mineral separation and to be the basis for advanced methods of mineral processing.

TPTTYKV, Here we show the first application of phage biotechnology to the processing of economically important minerals in ore slurries. A random heptapeptide library was screened for peptide sequences that bind selectively to the minerals sphalerite (ZnS) and chalcopyrite (CuFeS2). After several rounds of enrichment, cloned phage containing the surface peptide loops KPLLMGS and QPKGPKQ bound specifically to sphalerite. Phage containing the peptide loop TPTTYKV bound to both sphalerite and chalcopyrite. By using an enzyme-linked immunosorbant assay (ELISA), the phage was characterized as strong binders compared to wild-type phage. Specificity of binding was confirmed by immunochemical visualization of phage bound to mineral particles but not to silica (a waste mineral) or pyrite. The current study focused primarily on the isolation of ZnS-specific phage that could be utilized in the separation of sphalerite from silica. At mining sites where sphalerite and chalcopyrite are not found together in natural ores, the separation of sphalerite from silica would be an appropriate enrichment step. At mining sites where sphalerite and chalcopyrite do occur together, more specific phage would be required. This bacteriophage has the potential to be used in a more selective method of mineral separation and to be the basis for advanced methods of mineral processing.

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