Molecular Insight into the Binding Property and Mechanism of Sulfamethoxazole to Extracellular Proteins of Anammox Sludge.

Antibiotics are widely found in nitrogen-containing wastewater, which may affect the operation stability of anaerobic ammonium oxidation (anammox)-based biological treatment systems. Extracellular polymeric substances (EPSs) of anammox sludge play a pivotal role in combining with antibiotics; however, the exact role and how the structure of the leading component of EPSs (i.e., extracellular proteins) changes under antibiotic stress remain to be elucidated. Here, the interaction between sulfamethoxazole and the extracellular proteins of anammox sludge was investigated via multiple spectra and molecular simulation. Results showed that sulfamethoxazole statically quenched the fluorescent components of EPSs, and the quenching constant of the aromatic proteins was the largest, with a value of 1.73 × 10 ⁴ M ^-1 . The overall binding was an enthalpy-driven process, with Δ H = -75.15 kJ mol ^-1 , Δ S = -0.175 kJ mol ^-1 K ^-1 , and Δ G = -21.10 kJ mol ^-1 at 35 °C. The O-P-O and C═O groups responded first under the disturbance of sulfamethoxazole. Excessive sulfamethoxazole (20 mg L ^-1 ) would decrease the ratio of α-helix/(β-sheet + random coil) of extracellular proteins, resulting in a loose structure. Molecular docking and dynamic simulation revealed that extracellular proteins would provide abundant sites to bind with sulfamethoxazole, through hydrogen bond and Pi-Akyl hydrophobic interaction forces. Once sulfamethoxazole penetrates into the cell surface and combines with the transmembrane ammonium transport domain, it may inhibit the NH ₄ ⁺ transport. Our findings enhance the understanding on the interaction of extracellular proteins and sulfamethoxazole, which may be valuable for deciphering the response property of anammox sludge under the antibiotic stress.