No abstract

Biophysical characterization of type A botulinum neurotoxin (BoNT/A) complex along with its thermodynamic stability was assessed through a combination of various methods. BoNT/A exists as large complexes in association with neurotoxin associated proteins (NAPs). To evaluate its biophysical behavior, size-exclusion chromatography (SEC), multi-angled light scattering (MALS), enzyme linked immunosorbent assay (ELISA), and dynamic light scattering (DLS) were utilized. Initially, a single peak (peak 1) of SEC was observed at pH 6.0, and an additional peak (peak 2) appeared at pH 7.4 with a decrement of peak 1. Through MALS and ELISA, the peak 2 was determined to be BoNT/A dissociated from its complex. The dissociation was accelerated by time and temperature. At 37°C, dissociated BoNT/A self-associated at pH 7.4 in the presence of polysorbate 20. On the other hand, the dissociation was partly reversible when titrated back to pH 6.0. Overall, BoNT/A was more stable when associated with NAPs at pH 6.0 compared to its dissociated state at pH 7.4. The conventional analytical methods could be utilized to relatively quantify its amount in different formulations.

Biophysical characterization of type A botulinum neurotoxin (BoNT/A) complex along with its thermodynamic stability was assessed through a combination of various methods. BoNT/A exists as large complexes in association with neurotoxin associated proteins (NAPs). To evaluate its biophysical behavior, size-exclusion chromatography (SEC), multi-angled light scattering (MALS), enzyme linked immunosorbent assay (ELISA), and dynamic light scattering (DLS) were utilized. Initially, a single peak (peak 1) of SEC was observed at pH 6.0, and an additional peak (peak 2) appeared at pH 7.4 with a decrement of peak 1. Through MALS and ELISA, the peak 2 was determined to be BoNT/A dissociated from its complex. The dissociation was accelerated by time and temperature. At 37°C, dissociated BoNT/A self-associated at pH 7.4 in the presence of polysorbate 20. On the other hand, the dissociation was partly reversible when titrated back to pH 6.0. Overall, BoNT/A was more stable when associated with NAPs at pH 6.0 compared to its dissociated state at pH 7.4. The conventional analytical methods could be utilized to relatively quantify its amount in different formulations.