Wang Beibei, Xu Weijie, Wang Keke, Lin Hui, Zhang Xin, Ahmad Zulfiqar
College of Forest and Biotechnology, Zhejiang A & F University, Hangzhou, China.
Institute of Environment, Resource, Soil and Fertilizer, Zhejiang Academy of Agricultural Sciences, Hangzhou, PR China.
Microbiol Spectr. 2025 Sep 2;13(9):e0056225. doi: 10.1128/spectrum.00562-25. Epub 2025 Aug 12.
Current research on the microbial degradation of antibiotic residues primarily focuses on isolating degrading bacteria and characterizing their metabolites. The enzymatic mechanisms underlying these biotransformation processes remain poorly understood. Here, we investigated the molecular mechanisms involved in the biodegradation of oxytetracycline (OTC) by OTC-16 using integrated transcriptomic and proteomic sequencing analyses. OTC exposure significantly altered gene expression, identifying 1,158 and 863 differentially expressed genes on days 2 and 4, respectively. Proteomics revealed 352 and 818 differentially expressed proteins, predominantly involved in compound binding and redox processes. Integrated analysis identified (DNA oxidative demethylase), (urate oxidase), and (2-methylcitrate dehydratase) as key enzymes mediating OTC biodegradation through decarboxylation, carbon-carbon bond reduction, and dehydration reactions, confirmed by RT-qPCR. Protein-protein interactions subsequently highlighted supportive proteins, including glutathione S-transferase, ABC transporter ATP-binding protein, prpB, prpE, MarR regulators, and glyoxalase. Notably, and were successfully expressed in , and their OTC degradation-promoting activities were validated through assays. This study is the first to report the roles of and in antibiotic residue degradation, providing novel insights into the enzymatic mechanisms and laying a foundation for future genetic engineering and practical applications.IMPORTANCEOxytetracycline (OTC), a widely used antibiotic in agriculture and animal husbandry, frequently accumulates in the environment, posing significant ecological and human health risks by promoting antibiotic resistance. Understanding the microbial enzymatic pathways for OTC biodegradation is crucial for developing effective bioremediation strategies. This study provides a comprehensive molecular analysis of the OTC degradation process by OTC-16, identifying three key enzymes (, , and ) involved in OTC removal. Successfully expressing and in and validating their biodegradation activity further underscores their potential for biotechnological applications. These findings significantly enhance the knowledge of microbial antibiotic degradation mechanisms, offering practical molecular targets for environmental detoxification strategies.
目前关于抗生素残留微生物降解的研究主要集中在分离降解细菌并表征其代谢产物。这些生物转化过程背后的酶促机制仍知之甚少。在此，我们使用综合转录组学和蛋白质组学测序分析，研究了OTC - 16对土霉素（OTC）生物降解所涉及的分子机制。OTC暴露显著改变了基因表达，分别在第2天和第4天鉴定出1158个和863个差异表达基因。蛋白质组学揭示了352个和818个差异表达蛋白，主要参与化合物结合和氧化还原过程。综合分析确定了（DNA氧化脱甲基酶）、（尿酸氧化酶）和（2 - 甲基柠檬酸脱水酶）是通过脱羧、碳 - 碳键还原和脱水反应介导OTC生物降解的关键酶，这通过RT - qPCR得到证实。蛋白质 - 蛋白质相互作用随后突出了支持性蛋白质，包括谷胱甘肽S - 转移酶、ABC转运蛋白ATP结合蛋白、prpB、prpE、MarR调节因子和乙二醛酶。值得注意的是，和在中成功表达，并且它们促进OTC降解的活性通过测定得到验证。本研究首次报道了和在抗生素残留降解中的作用，为酶促机制提供了新的见解，并为未来的基因工程和实际应用奠定了基础。重要性土霉素（OTC）是农牧业中广泛使用的抗生素，经常在环境中积累，通过促进抗生素耐药性对生态和人类健康构成重大风险。了解OTC生物降解的微生物酶促途径对于制定有效的生物修复策略至关重要。本研究对OTC - 16降解OTC的过程进行了全面的分子分析，确定了参与OTC去除的三种关键酶（、和）。在中成功表达和并验证它们的生物降解活性进一步强调了它们在生物技术应用中的潜力。这些发现显著增强了对微生物抗生素降解机制的认识，为环境解毒策略提供了实际的分子靶点。
