Synthetic biology platform targets antibiotic residues in water
image: Graphical abstract
Credit: Hao Ren, Meilin Qin, Lin Zhang, Zemiao Li, Yuze Li, Qian He, Jiahao Zhong, Donghao Zhao, Xinlei Lian, Hongxia Jiang, Xiaoping Liao, Jian Sun
Researchers at South China Agricultural University have developed a novel synthetic biology-based platform to degrade tetracycline residues in aquatic environments. The study, published in Engineering, details the creation of a modular enzyme assembly called FerTiG, which integrates multiple functional modules to efficiently remove tetracycline (TC) from various water sources.
The FerTiG system is inspired by the microcompartment structures found in living organisms, which compartmentalize enzymes to enhance their stability and efficiency. In this engineered construct, the TC-degrading enzyme Tet(X4) is combined with a glucose dehydrogenase (GDH) module for cofactor recycling and encapsulated within a ferritin cage for protection. This design aims to address the challenges of using standalone enzymes, which often require expensive cofactors and are susceptible to environmental degradation.
The construction of FerTiG involved genetic fusion of Tet(X4) with GDH and recombinant human heavy-chain ferritin. The resulting complex was expressed in E. coli and purified using a HisTrap column. Characterization of FerTiG revealed a globular structure with an increased hydrodynamic radius compared to free Tet(X4), indicating successful encapsulation within the ferritin cage.
The GDH module in FerTiG is designed to regenerate the NADPH cofactor using glucose, thereby reducing the need for external cofactor supplementation. This feature was confirmed through colorimetric assays, which showed that NADPH regeneration occurred only in the presence of both GDH and glucose. The incorporation of GDH significantly enhanced the efficiency of TC degradation, as demonstrated in microbiological inhibition assays using Bacillus stearothermophilus. FerTiG was able to completely remove TC from the solution within a shorter time frame compared to free Tet(X4).
The ferritin cage provides FerTiG with resilience against environmental stresses. Experiments showed that FerTiG retained its activity under extreme conditions, such as high ionic strength, low or high pH, and prolonged UV exposure, where free Tet(X4) lost its functionality. Additionally, FerTiG demonstrated stable storage properties, maintaining its degradation efficiency over a week at room temperature and 4 °C, unlike free Tet(X4), which lost activity within a day at 37 °C.
To assess the practical application of FerTiG, the researchers tested its performance in different water matrices, including tap water, lake water, livestock sewage, and pharmaceutical wastewater. FerTiG effectively removed TC residues from all tested matrices, achieving complete removal in lake water and livestock sewage within 3 hours. The degradation products were analyzed using high-performance liquid chromatography (HPLC) and quadrupole time-of-flight mass spectrometry (QTOF-MS), revealing a series of transformation intermediates that indicated the breakdown of TC into smaller, potentially biodegradable molecules.
The study also evaluated the biosafety of FerTiG. Predictive toxicity analysis using the ECOSAR program suggested that the transformation products had low aquatic toxicity. Furthermore, zebrafish embryos exposed to FerTiG showed no morphological abnormalities or delayed hatching, indicating the ecological safety of the system. In vivo biosafety tests in mice demonstrated that FerTiG did not cause significant changes in body weight or histological damage to vital organs such as the liver, kidneys, and intestines.
The development of FerTiG represents a step towards sustainable and eco-friendly solutions for managing antibiotic residues in water. By combining the benefits of enzyme-based and living organism-based approaches, this modular system offers a promising strategy for environmental remediation. Future work may focus on scaling up the production of FerTiG and exploring its application in treating other types of antibiotic residues.
The paper “Modular Engineering of a Synthetic Biology-Based Platform for Sustainable Bioremediation of Residual Antibiotics in Aquatic Environments,” is authored by Hao Ren, Meilin Qin, Lin Zhang, Zemiao Li, Yuze Li, Qian He, Jiahao Zhong, Donghao Zhao, Xinlei Lian, Hongxia Jiang, Xiaoping Liao, Jian Sun. Full text of the open access paper: https://doi.org/10.1016/j.eng.2025.03.033. For more information about Engineering, visit the website at https://www.sciencedirect.com/journal/engineering.