Goodbye Azodye UCL iGEM 2014

About Our Project

The Problem: Azo Dyes in the environment

Since their accidental discovery by Sir William Henry Perkin in 1853, azo dyes have become one of the most popular forms of synthetic colourant. These dyes are currently used in the industrial manufacture of a variety of products, ranging from clothing and upholstery to cosmetics and tattoo ink, as well as many others.

Although azo-dyes are widely regarded as a safe and stable form of synthetic colourant, some of them can take on dangerous properties after they have been broken down by enzymes in the guts of organisms.

In the textile industry alone, the global annual production of dyes amounts to a million metric tons. In many countries, the leftover dye effluent produced by industrial manufacturers is often not properly disposed of, or removed, during water treatment.

This results in the accumulation of azo dyes in water bodies where they are then ingested by aquatic organisms. Additionally, irrigation of agricultural lands with dye polluted water severely affects soil fertility and plant growth.

The products of this enzymatic breakdown have been found to be both mutagenic and carcinogenic, and have been linked to increased occurrences of several different forms of cancer if they enter the food chain. Despite this toxicity and it's potential effect on human health, little to no effort has been made to dispose of these leftover azo dyes more responsibly.

As a result, development of remediation technologies for treatment of dye containing waste waters has been a matter of major concern for environmentalists.

The Solution: GoodBye Azo Dye, UCL iGEM 2014

We have created an Azo-Remediation Chassis (ARC), a complete synthetic azo dye decolourising device in E. coli. The ARC harnesses several different independent enzymes that degrade azo dyes and their breakdown products. This allows the development of a bioengineered process preventing accumulation of carcinogenic azo dye products in industrial wastewater.

The diagram above demonstrates how our E. Coli plasmid incorporates the following enzymes: Azoreductase from Pseudomonas Aeruginosa, Lignin Peroxidase from Phanaerochaete Chrysosporium, Azoreductase & Bacterial Peroxidase from Pseudomonas Putida, and Bacterial Peroxidase & Laccase from Bacillus Subtilis.

Azoreductase from Pseudomonas Aeruginosa

Azoreductase from Pseudomonas aeruginosa is intended to work complementary with azo dyes, in order to cover a wider spectrum of dyes more efficiently.

The reaction pathway on the right demonstrates how azoreductase will cleave the azo-bond (N=N), producing aromatic amines. However, these amines are highly toxic; hence we have incorporated further enzymes into our ARC.

Lignin Peroxidase from Phanaerochaete Chrysosporium

Aromatic amines produced by azoreductase cleavage are oxidised, resulting generally in a substitution of the amine groups by other oxygenated groups like carboxyls or carbonyls.

In this case, the oxidation will be carried out by Lignin Peroxidase from Phanaerochaete Chrysosporium. The mechanisms for these oxidations vary from dye to dye, hence we have incorporated laccase and other peroxidases into our ARC.

Azoreductase & Bacterial Peroxidase from Pseudomonas Putida

Azoreductase from Pseudomonas Putida performs the cleavage of the azo-bond (N=N), working in conjunction with Azoreductase from Pseudomonas aeruginosa - maximising the efficiency of our ARC.

Bacterial Peroxidase from Pseudomonas Putida performs the oxidisation of the aromatic amines, working in conjunction with Lignin Peroxidase from Phanaerochaete Chrysosporium.

Bacterial Peroxidase & Laccase from Bacillus Subtilis

Bacterial Peroxidase & Laccase from Bacillus Subtilis performs the oxidisation of the aromatic amines, working in conjunction with Lignin Peroxidase from Phanaerochaete Chrysosporium and Bacterial Peroxidase from Pseudomonas Putida - maximising the efficiency of our ARC.

A bioprocess employing the ARC in an industrial setting has been developed and various modes of operation explored. This may serve as an end-of-pipe, lucrative addition to facilities expelling azo dye contaminants. Furthermore, xenobiological approaches to biosafety are considered and a proposal for an “azotrophic” organism paves the way for a new era in synthetic biology biosafety.

For our iGEM project we developed a process to controllably degrade and detoxify the excess azo dye effluent at the source - the textile factories - before they even reach the water systems. We achieved this goal by introducing the genes for three enzymes related to the degradation of these dyes: azoreductase, laccase, and lignin peroxidase into a host E.coli cell to create an enhanced azo dye decolourising organism.

We also designed an integrated end-of-pipe method for detoxifying dye factory wastewater effluent streams by incorporating our engineered E. coli strain in a two-stage process to ensure optimal conditions for the degradation of azo dyes within a batch bioreactor system. The potential for scalability of this method would present various economic and environmental advantages for industries that generate large amounts of dyestuff. The system we have developed could also be enhanced to become a modular bioprocess method for wastewater treatment of other toxic, normally recalcitrant chemicals.

This year, UCL has a highly interdisciplinary team of undergraduates and postgraduates, forming a synbio community at UCL. We are all genuinely delighted to be trying to bring synthetic biology to the world around us. This year we have accomplished immense public engagement and tackled key issues regarding policy and practices.

Contact Us

University College London
Gower Street - London
Biochemical Engineering Department
Phone: +44 (0)20 7679 2000

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