Regulation of Botulinum neurotoxin half‑life through the ubiquitin‑proteasome system

Botulinum neurotoxin (BoNT), renowned for its potency, is used in both therapeutic and cosmetic applications; however, it is constrained by a relatively short half‑life and limited duration of efficacy. The present study investigated the role of the ubiquitin‑proteasome system in BoNT degradation and evaluated strategies to extend its half‑life by targeting specific lysine residues. Ubiquitination was observed in the light chains of BoNT/A1 (K335 and K417), BoNT/A2 (K335) and BoNT/E (K62 and K288), which were identified as key ubiquitination sites. The substitution of these lysine residues to arginine inhibited ubiquitination, improving protein stability and extending the half‑life of BoNT. These findings offer a potential strategy to enhance the therapeutic efficacy of BoNT by prolonging its stability, paving the way for future advancements in BoNT‑based treatments.