Painkillers such as diclofenac are often used in pharmacological ointments because of their anti-inflammatory and analgesic properties, as well as the advantage of local use, making it exceedingly popular among doctors and patients.
Diclofenac and other micropollutants accumulate in wastewater due to overuse, thus contaminating the environment. The current treatment in sewage treatment plants is expensive and complex, however our vision is to make this process efficient and cost effective through the use of genetic engineering.
Our project involves the modification of the green algae Chlamydomonas reinhardtii, enabling the chemical decomposition of diclofenac and leading to its functional degradation.
To achieve this, we will clone genes from two enzymes into the genome of our primary organism, the green algae Chlamydomonas reinhardtii, as well as into the bacterium Escherichia coli as a control. These enzymes are two different laccases. Laccases are oxidases that have a multicopper center. This allows them to oxidize their substrates, taking up an electron and transferring it to O2. Water is produced as a by-product. (Kittl et al., 2012; Zerva et al., 2019)
Our goal is to achieve a high production of laccase in our organisms and then be secreted to the extracellular medium, i.e. the wastewater of the sewage treatment plants. The enzymes must withstand the prevailing conditions in the sewage treatment plant, such as a certain pH and temperature range, to be able to begin oxidation.
We will test the activity of the enzymes on substances such as ABTS and then investigate the degradation of the painkiller diclofenac. There is even the possibility that laccases can oxidize a whole range of micropollutants that are in high concentrations present in the wastewater and difficult to remove by conventional processes, e.g. antidepressants, antiepileptics and beta-blockers. (Bilal et al., 2019)
The problem with these micropollutants is their fortification in wastewater due to incomplete degradation. Among the substances that are highly toxic to aquatic life are diclofenac, ibuprofen and paracetamol, some of which occasionally can also accumulate in animals, where they still have unexplored effects on hormonal mechanisms of action. (Bilal et al., 2019)
Because we would like to prevent contamination of wastewater with genetically modified organisms (GMOs), and in order to achieve the best possible growth of green algae crops, they are to be bred in a bioreactor outside the wastewater treatment plants. The laccases enter the wastewater basin via a filter system that generates only the passage of the enzymes, but not the passage of the laccase laden organisms.
It is our vision to create a cost-effective and efficient approach to a cleaner and healthier environment.
Bilal, M. et al. (2019) ‘Emerging contaminants of high concern and their enzyme-assisted biodegradation – A review’, Environment International, 124, pp. 336–353. doi: 10.1016/j.envint.2019.01.011.
Kittl, R. et al. (2012) ‘A chloride tolerant laccase from the plant pathogen ascomycete Botrytis aclada expressed at high levels in Pichia pastoris’, Journal of Biotechnology, 157(2), pp. 304–314. doi: 10.1016/j.jbiotec.2011.11.021.
Zerva, A. et al. (2019) ‘Applications of Microbial Laccases: Patent Review of the Past Decade (2009–2019)’, Catalysts. Multidisciplinary Digital Publishing Institute, 9(12), p. 1023. doi: 10.3390/catal9121023.