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Team:Clemson/Project
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Abstract
Polychlorinated biphenyls (PCBs) are widespread, cancer-causing pollutants left-over mainly from manufacture of capacitors and electric motors. There are over 200 possible PCBs, derivatives of biphenyl, which share the same biodegradation pathways in bacteria. Our team is using a genetic engineering approach to produce a small consortium of E. coli that should efficiently degrade biphenyl, and it is hoped that this same system can be adapted for the bioremediation of PCBs. Natural bioremediation by native bacterial communities is exceedingly slow due to the recalcitrant nature of PCBs and their hydrophobic properties which reduce the bioavailability to potential catabolizers. We are taking a three-pronged approach in an attempt to increase the efficiency of biphenyl bioremediation—attraction of biphenyl-degrading E. coli by other guiding bacteria, overexpression of the biphenyl catabolic enzymes, and production of a biosurfactant to increase the solubility of biphenyl. Together, this system should significantly increase the rate of biphenyl degradation. Motivation
From 1955 to 1987 in Pickens, South Carolina (the neighboring town to Clemson, SC) Sangamo-Weston Inc. had multiple factories, where they built capacitors. The dielectric fluid used in these capacitors contained Polychlorinated Biphenyls (PCBs). Due to improper land-burial of off-specification capacitors and wastewater treatment sludges that were built both on-site and off-site, PCBs contaminated the water supply of Lake Hartwell and its tributaries. Since PCBs have low water solubility, they remain mostly in the organic sediments and as particulate matter. Furthermore, PCB sediment concentrations are within the range of injury for benthic macroinvertabrates and exceed healthy levels in the fish of Lake Hartwell – an 87.5 square mile recreational lake that serves both South Carolina and Georgia areas. Introduction
Overall, our project goal is to create a three-component bacterial system: signaling, biosurfactant, and PCB degradation. First is an expansion of the UT-Tokyo 2011 Team’s bioremediation specific system, consisting of guider and worker bacteria. The system (currently activated by UV light) causes the guider bacteria to produce Aspartate A, which is converted to L-Asp, a natural chemoattractant for E. coli. Thus, both guiders and workers will respond when the bacteria come into contact with the substrate. However, the reaction producing L-Asp is reversible and would cause the bacteria to no longer stay with the substrate. In order to circumvent this, UT-Tokyo created a system with a chromosomal cheZ knockout E. coli stain and initiated substrate-dependent cheZ expression from a plasmid. Thus, in the presence of the substrate, the lambda phage repressor cI is produced, binding to the cI-repressed promoter, inhibiting cheZ expression, and halting bacterial movement. In order for this system to work for our purposes, we are to replace the UV light promoter with a PCB specific promoter. Ideally we would like to implement this system in magnetotactic bacteria since PCBs are insoluble particles that remain in the sediments of Lake Hartwell, we will need the guiders to dive to the bottom. The workers in this system will be comprised of two genetically different bacteria: one programmed to produce a biosurfactant and the other to degrade the PCBs to less harmful byproducts.
Overview of Project Approach
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