20.500.12556/RUL-182732 A deep dive into spin-labeled polysorbate’s interaction with therapeutic antibody using 2D NMR, EPR and MD simulations Polysorbates (PS) are widely used surfactants in biopharmaceutical formulations playing a crucial role in protecting proteins against mechanical stress and interface-induced damage. However, their susceptibility to degradation can compromise their function and lead to particle formation. Recent studies suggest that monoclonal antibodies (mAbs) may mitigate PS degradation catalyzed by histidine chloride buffer, indicating the presence of protein-PS interactions. In this study, we investigated these interactions using NMR, starting with $^1$H T$_2$ CPMG filter experiments and methyl fingerprinting, which failed to detect interactions. To enhance sensitivity, we synthesized spin-labeled PS (SLPS), enabling paramagnetic relaxation enhancement (PRE) NMR experiments, specifically amide fingerprinting, which successfully revealed interactions. Complementary electron paramagnetic resonance (EPR) measurements of SLPS and mAb also detected interactions, but only in the presence of sucrose, underscoring their weak and transient nature. Additionally, molecular dynamics simulations identified potential interaction hotspots on the antibody structure, providing mechanistic insights into these interactions. monoclonal antibody interaction polysorbate nuclear magnetic resonance electron paramagnetic resonance molecular dynamics monoklonsko protitelo interakcija polisorbat jedrska magnetna resonanca elektronska paramagnetna resonanca molekularna dinamika farmacevtska kemija biokemija zdravila true false true Angleški jezik Slovenski jezik Članek v reviji 2026-05-21 15:04:35 2026-05-21 15:04:43 2026-05-22 04:49:56 0000-00-00 00:00:00 2025 0 0 9 str. art. 114892 Vol. 217 Dec. 2025 0000-00-00 Zaloznikova Objavljeno NiDoloceno 0000-00-00 0000-00-00 0000-00-00 577:615.32 1873-3441 10.1016/j.ejpb.2025.114892 255171075 1-s2.0-S0939641125002693-main.pdf 1-s2.0-S0939641125002693-main.pdf 1 D4EA6A4D57E4448F1E55337DC02D5D01 a3f10cd1172ff5f1c5561d82488cb58af50e60edac9a738992fb1ea1a05ae6c3 b61e7654-5515-11f1-b0ab-0050569b8976 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=234249 https://www.sciencedirect.com/science/article/pii/S0939641125002693 0 528f9e81-5515-11f1-b0ab-0050569b8976 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=234248 Fakulteta za farmacijo 0 0 0