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UCONN iGEM




Thallium Overview

Thallium (TI) is an uncommon, but highly toxic element to mammals. Thallium has forty-seven isotopes having atomic masses between 179 and 210 grams, and all isotopes of thallium are toxic.

Thallium is a very prominent pollutant and spreads through the environment as waste from drilling in sulfur deposits. The United States’ accepted maximum contaminant value for thallium in drinking water is 5 μg/L. In dig sites, ores are extracted through heap leaching, which places mixtures of ores on a liner and adds chemicals as separating factors to the ores. The target ore, often sulfur in cases of thallium pollution, is extracted and the excess ores are replaced back into sediment beds. As water also flows through these sediment beds, thallium is added to the runoff.

Humans take up significant amounts of TI either directly through drinking water or absorbed through the skin and mucous membranes. Once inside the body, thallium becomes deposited in bones and the renal medulla. Symptoms of thallium poisoning include hair loss, changes in blood pH, decreased kidney function and intestinal and liver problems. The maximum contaminant level (MCL) is the highest concentration that is allowed in drinking water, when considering current methods of removal. The current MCL of thallium is 2 ppb. The EPA cannot enforce any contamination levels below 2 ppb because current water treatment protocols cannot remove these trace amounts of thallium. The maximum contaminant level goal (MCLG), however, is 0.5ppb. The MCLG is the amount of contaminant in drinking water that will cause no negative health effects. The estimated average concentration of thallium in drinking water is 7.2 ppb, almost fifteen times the MCLG. Currently, the only methods approved by the EPA for thallium removal are activated alumina and ion exchange. Unfortunately, activated alumina and ion exchange are not necessarily the most effective techniques for thallium removal.

Activated alumina is a highly porous material that is created by dehydroxylating aluminum hydroxide. Depending on pH, various ions can adsorb onto the activated alumina. Activated alumina devices have been able to lower thallium levels below 2 ppb. However, various constraints hinder its ability to effectively remove thallium. Suspended solids in water can obstruct the activated alumina, and the washes required to unsaturate the activated alumina device are detrimental to the ion removal efficiency. Alternatively, activated alumina devices can be replaced upon saturation, but disposal of used activated alumina can be hazardous, depending on what is bound to the activated alumina. Furthermore, storing activated alumina devices as replacements is not feasible for long periods of time. The activated alumina particles cement together when left unused for several days. As a result, only short-term storage of the device is feasible. Ion exchange columns are an alternative to activated alumina.

Ion exchange columns contain resin beads which store positive or negative ions for anion or cation exchange, respectively. Due to the presence of positively or negatively charged species on the beads, ions of opposite charge bind to the resin beads through electrostatic interactions. Ion exchange columns, as with the activated alumina, are able to lower thallium levels below 2ppb. However, ion exchange columns, like the activated alumina, face the same problem: Suspended particles can accumulate on the resin, hindering its ability to effectively interact with ions in the water15. Suspended particles are prevalent in water. A majority of particles in wastewater are approximately 40-50um in diameter. River water contains many minerals and organic matter in the form of loose sediment. Drinking water contains small amounts of solid particles that range from “colloidal sizes to 100um”. In summary, water samples for purification will most likely contain species which can hinder the ion exchange column. After considering the disadvantages of both the activated alumina particles and ion exchange columns, one can see that an improved method of thallium removal is needed.

The radioactive isotope Tl-201(ionic form: Tl+) was first implicated for clinical use by Lebowitz, et al. 1976. Scintigraphy is a form of nuclear medicine in which a radioisotope is administered to the patient at low, nontoxic doses. In cardiac scintigraphy, Lebowitz et al discovered that Tl-201 can be used as part of diagnostic stress test, now known as the thallium stress test. The thallium stress test allows clinicians to perform non-invasive myocardial screenings. Tl-201 is used because, in the blood, Tl-201 is in its ionic form, Tl+, which acts as a potassium (K+) analog, allowing Tl-201 uptake by cardiac cells’ Na/K-ATPases.. The utility of the Tl-201 in myocardial scintigraphy as a potassium analog indicates that similar potassium transport systems in bacteria can potentially be used to facilitate uptake of thallium.





Trk Proteins

TrkA is the primary protein in the Trk family. TrkG and TrkH are membrane proteins that help TrkA bind to the inside of the cellular membrane. However, in order to have Trk function, only TrkG or TrkH must be present, not both. The TrkG gene can be deleted without having much influence on Trk function, and is therefore likely to have a similar function to either TrkH or TrkE. TrkE is a membrane protein but unlike TrkH and TrkG it does not help bind TrkA to the membrane. Through exploratory processes, scientists have suggested that TrkE and TrkG have similar but not identical functions.

TrkH or trkG span the cell membrane, while trkA, connected to one of the previous proteins within the cytoplasm of the cell, serves as a peripheral NAD(H) binding protein. Within the trkH system, trkE, carrying ATP, interacts with the rest of the transport protein system to stimulate transport activity. While the explicit details of the interaction between these three proteins remain unknown, it has been hypothesized that ATP is transferred from trkE to trkA, causing trkA to shift to a more open form, resulting in dilation within the upper layer of RCK domains within trkA. This dilation pulls on the interconnecting helices between the attached trkA and trkH, opening the intercellular gate that permits the flow of K+ ions into or out of the cell.

trkA: This protein binds with NAD+ to create the driving force to move K+ ions across the membrane. (Schlosser et al 1993). This is partially dependent on the proton gradient. Mutations in the TrkA inhibits the Trk complex from transporting K+, indicating that it is necessary for proper protein function. TrkA also is involved in regulation of the TrkH subunit’s ion channel (Rhoads, Waters, & Epstein 1976). The structure of TrkA consists of 4 protomers, creating a symmetric gating ring. The ATP binding site is located on the Arg-100 residue, which is necessary to regulate the TrkH ion channel. There are eight total Regulate-the-Conductance-of-K+ (RCK) domains that are involved in the NADH binding (Cao et al 2013).

trkE (sapD): Many studies refer to trkE as sapD, as the sapABCDF operon contains the trkE gene locus, putatively at sapD. The TrkE protein binds ATP and is thought to be the activator of the Trk potassium uptake system. Only ATP binding is necessary for Trk system activity, hydrolysis is nonessential (Stewart et al., 1985). The crystal structures for this protein have not been determined experimentally, although several models have been proposed. (http://www.uniprot.org/uniprot/P0AAH4). TrkE expression is necessary for trkH activity and increases the rate of the TrkG system.

trkG: This subunit is part of the transmembrane component in the Trk system. It works analogously to the TrkH subunit. Compared to other variants, TrkG provided a faster rate of ion transport. In addition to its affinity for K+, it also showed it could uptake Rb at a slower rate (Schlösser et al. 1995). The crystal structure has not been determined yet, but according tto UniProt (http://www.uniprot.org/uniprot/P23849), four binding sites have affinity for K+ ions (and Rb ions) Only either TrkG or TrkH is requried for function of the Trk system (Johnson et al. 2009).

trkH: This protein is required for the uptake of K+ ions are taken in through the cell membrane. It is permeable to other ions such as Na+, Li+, and Rb+. The TrkH protein is necessary for bacterial growth in low K+ environments (Cao et. al., 2011). TrkH assembles with TrkA (NAD binding protein), and requires the TrkE protein to be functional (Bossemeyer, 1989). K+ uptake is dependent on intracellular [NAD] and may be linked to H+ symport (Rhoads and Epstein, 1977). TrkH is a transmembrane protein consisting of two TrkH protomers , and each protomer contains an ion permeation pathway. With respect to a plane perpendicular to the membrane, the TrkH protomers would display twofold symmetry and form the shape of a parallelogram. Each protomer is composed of five domains labelled D0 through D4. D0 is composed of two transmembrane segments while the other four domains resemble K+ channels. A highly conserved Arg468 residue forms a constriction with the intramembrane loop, which is required for K+ to pass. Arg468 substitution increases K+ permeation significantly, similar to various deletions of the intramembrane loop (Cao et. al., 2011).

    Reference List

  1. Bossemeyer, D., Borchard, A., Dosch, D.C., Helmer, G.C., Epstein, W., Booth, I.R., and Bakker, E.P. (1989). K+-transport Protein TrkA of Escherichia coli Is a Peripheral Membrane Protein That Requires Other trk Gene Products for Attachment to the Cytoplasmic Membran. The Journal of Biological Chemistry 264, 16403-16410.

  2. Cao, Y., Jin, X., Derebe, M., Levin, E., Kabaleeswaran, V., Pan, Y., Punta, M., Love, J., Weng, J., Quick, M., et al. (2011). Crystal structure of a potassium ion transporter, TrkH. Nature 471, 336-340.

  3. Cao, Y., Pan, Y., Huang, H., Jin, X., Levin, E.J., Kloss, B., and Zhou, M. (2013). Gating of the TrkH ion channel by its associated RCK protein TrkA. Nature 496, 317-322.

  4. Gomez-Gonzalez, M.A., Garcia-Guineaa, J., Labordab, F., and Garrido, F. (2015). Thallium occurrence and partitioning in soils and sediments affected by mining activities in Madrid province (Spain). Science of the Total Environment 536, 268-278.

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  13. Schlosser, A., Hamann, A., Bossemeyer, D., Schneider, E., and Bakker, E.P. (1993). NAD+ binding to the Escherichia coli K+-uptake protein TrkA and sequence similarity between TrkA and domains of a family of dehydrogenases suggest a role for NAD+ in bacterial transport. Molecular Microbiology 9, 533.

  14. Stewart, L.M.D., Bakker, E.P., and Booth, I.R. (1985). Energy Coupling to K+ Uptake Via the Trk System in Eschevichiu colk the Role of ATP. Journal of General Microbiology 131, 77-85.

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