Fig. 4
From: Elucidating the activation mechanism of botulinum neurotoxin a: role of α-clostripain and NTNH
Protection by NTNH is essential for specific BoNT/A activation by α-clostripain. (A) SDS-PAGE analysis for evaluating the activity of α-clostripain and C. botulinum A culture supernatant on rBoNT/Ai in the presence or absence of rNTNH. Lane 1–MW marker; Lane 2—rBoNT/Ai; Lane 3–rNTNH; Lane 4 – rBoNT/Ai + rNTNH + rα-clostripain; Lane 5–rBoNT/Ai + rα-clostripain; Lane 6—rBoNT/Ai + rNTNH + C. botulinum A culture supernatant; Lane 7—rBoNT/Ai + C. botulinum A culture supernatant. All reactions were performed at pH 6.0. (B) The protection of BoNT/A by NTNH is pH-dependent and influenced by the BoNT/A-NTNH interaction, which associate under acidic pH and dissociate under alkaline pH. Lane 1–MW marker; Lane 2–rBoNT/Ai; Lane 3–rNTNH; Lane 4—rBoNT/Ai + rNTNH + rα-clostripain at pH 6.0; Lane 5—rBoNT/Ai + rNTNH + rα-clostripain at pH 8.0; Lane 6—rBoNT/Ai + rNTNH + C. botulinum A culture supernatant at pH 6.0; Lane 7—rBoNT/Ai + rNTNH + C. botulinum A culture supernatant at pH 8.0. The 100-kDa heavy chain of rBoNT/Ai is marked with * and the 50-kDa light chain of rBoNT/A is marked with #.
