Team:SNU India/Description


Project Inspiration and Description

Endocrine disrupting compounds (EDC’s): an emerging public health and environmental threat.
Since the inception of global industrialization, steroidal estrogens have become an emerging and serious concern. Worldwide, steroid estrogens including estrone, estradiol and estriol, pose serious threats to soil, plants, water resources and humans [Adeel et.al, 2017]. Once in the environment, estrogens are taken up and accumulated by plants [Zheng et.al, 2014], which finally makes its way into human system. Similarly, estrogens also affect several aquatic populations such as fish, where endocrine responses can lead to a skewed sex ultimately leading to a decline in their populations [Scholz and Kluver, 2009]. Estrogen also has been shown to play a significant role in many public health issues, including incidence of breast cancer in women, prostate cancer in men, early-onset of puberty and other hormonal disorders [Adeel et.al, 2017]. Besides natural estrogen, there are other phenolic molecules which mimic the activity of estrogen, such as Bisphenol A (Plastic waste), EE2 (contraceptive pills), and synthetic Estrone (E1). Currently, there are various technologies such as treatment with UV, ozonation, or adsorption by activated carbon, and bioreactor-membrane filters, used for removal of EDCs and other micro-pollutants from wastewater. However, many such technologies are not widely utilized, due to them being not that effective, technical nature and financial burden [Xiong et,al, 2018].
Laccases: Enzymes for removing EDCs and various phenolic pollutants
Laccases are multi-copper ligno-lytic oxidoreductases, having the potential to efficiently degrade EDCs [Watkins and Nicell, 2011] (Fig 1). Many systems such as backed or fixed bed reactors use immobilized laccase columns, live fungi-based treatment systems and immobilized laccase-beads were developed for degradation of estrogens and many other phenolic pollutants from wastewater [Becker et.al, 2017]. One of the major limiting factors for scaling up the process is purification and immobilization of the enzyme, and requirement of mediators that can increase the efficiency of laccase activity on substrates with lower redox potential [Becker et.al, 2017].
Microalgae: A wastewater treatment solution

Microalgae provide an attractive solution for removal of various macro and micro-pollutants from effluent wastewater [Xiong et,al, 2018; Raouf et.al, 2012]. They are photosynthetic microorganisms, which can utilize inorganic nitrogen and phosphorus, adsorb, absorb and remove organic micropollutants, heavy metals.Microalgae accumulation will yield biomass in the process, which can have several implications such as biofuel production, fertilizers, nutraceuticals and pharmaceuticals.


Figure 1: Laccase mediated degradation, where T1, T2, T3 and T4 are the four copper atoms present in the reaction centre of the enzyme laccase and (+2) is the oxidation state of the copper atoms, before reaction and (+1) is the oxidation state of the copper atoms after the reaction. Microalgae-bacterial consortium have several synergistic interactions that can be harnessed for biological production of laccases and its mediators for EDC degradation, wastewater bioremediation, and biofuel production.


In addition, this approach can also be used for bioremediation of other phenol-based dyes used extensively in textile industry, and for lignin degradation derived from the paper and pulp industry along with pharmaceutical waste degradation for degradation of poly-phenolic drugs.

AlBaCo: Integrated Detection, quantification and degradation

Initially we had planned our system to be a consortium of two types of transformed bacteria and an untransformed microalga. One bacteria was to be engineered to contain a reporter module, while the other bacteria with a bacterial laccase to enable EDC degradation. Our reporter module consisted of a reporter protein i.e., RFP and downstream to it an enzyme (TAL), which would produce a laccase mediator to enhance laccase activity that would be expressed only in presence of estrogen/EDCs.

However, as mentioned above, fungal laccases are known to be more efficient as compared to bacterial laccases when it comes to EDC degradation. This motivated us to ideate a consortium of a transformed microalgae and a transformed bacteria. The engineered bacteria would consist of the reporter module, while the microalgae would be engineered to secrete a fungal laccase for EDC degradation.

References:
  1. Adeel, M., Song, X., Wang, Y., Francis, D., & Yang, Y. (2017). Environmental impact of estrogens on human, animal and plant life: A critical review. Environment International, 99, 107–119. https://doi.org/10.1016/j.envint.2016.12.010
  2. Zheng, W., Wiles, K. N., Holm, N., Deppe, N. A., & Shipley, C. R. (2014). Uptake, Translocation, and Accumulation of Pharmaceutical and Hormone Contaminants in Vegetables. In Retention, Uptake, and Translocation of Agrochemicals in Plants (pp. 167–181). American Chemical Society. https://doi.org/10.1021/bk-2014-1171.ch009
  3. Scholz, S., & Klüver, N. (2009). Effects of Endocrine Disrupters on Sexual, Gonadal Development in Fish. Sexual Development, 3(2–3), 136–151. https://doi.org/10.1159/000223078
  4. Xiong, J.-Q., Kurade, M. B., & Jeon, B.-H. (2018). Can Microalgae Remove Pharmaceutical Contaminants from Water? Trends in Biotechnology, 36(1), 30–44. https://doi.org/10.1016/j.tibtech.2017.09.003
  5. Cardinal-Watkins, C., & Nicell, J. A. (2011). Enzyme-Catalyzed Oxidation of 17β-Estradiol Using Immobilized Laccase fromTrametes versicolor. Enzyme Research, 2011, 1–11. https://doi.org/10.4061/2011/725172
  6. Becker, D., Rodriguez-Mozaz, S., Insa, S., Schoevaart, R., Barceló, D., de Cazes, M., … Wagner, M. (2017). Removal of Endocrine Disrupting Chemicals in Wastewater by Enzymatic Treatment with Fungal Laccases. Organic Process Research & Development, 21(4), 480–491. https://doi.org/10.1021/acs.oprd.6b00361
  7. Abdel-Raouf, N., Al-Homaidan, A. A., & Ibraheem, I. B. M. (2012). Microalgae and wastewater treatment. Saudi Journal of Biological Sciences, 19(3), 257–275. https://doi.org/10.1016/j.sjbs.2012.04.005
  8. Park, S., Lee, Y., Lee, J.-H., & Jin, E. (2013). Expression of the high light-inducible Dunaliella LIP promoter in Chlamydomonas reinhardtii. Planta, 238(6), 1147–1156. https://doi.org/10.1007/s00425-013-1955-4