Bifunctional Inhibition of Botulinum Neurotoxin A Protease: Unexpected Active Site Inhibition Enhances Covalent Targeting of an Allosteric Site

Botulinum neurotoxin (BoNT) is the most toxic protein known to man and a Tier 1 bioterrorism agent. Among its serotypes, BoNT/A possesses the greatest potency and persistence, as such strategies to counteract it are highly coveted. Bifunctional molecules incorporating both metal chelation and a cova...

وصف كامل

محفوظ في:
التفاصيل البيبلوغرافية
المؤلف الرئيسي: Mingliang Lin (3745552) (author)
مؤلفون آخرون: Nishant Karadkhelkar (21083917) (author), Yanjie Li (1463440) (author), Lisa M. Eubanks (1371738) (author), William H. Tepp (226108) (author), Sabine Pellett (528430) (author), Kim D. Janda (163029) (author)
منشور في: 2025
الموضوعات:
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الوصف
الملخص:Botulinum neurotoxin (BoNT) is the most toxic protein known to man and a Tier 1 bioterrorism agent. Among its serotypes, BoNT/A possesses the greatest potency and persistence, as such strategies to counteract it are highly coveted. Bifunctional molecules incorporating both metal chelation and a covalent warhead have shown great potential for blunting BoNT/A LC’s toxicity/longevity. To further explore this idea, new warheads as well as zinc metal-chelating scaffolds were prepared and examined. The structure–activity relationship and kinetic analyses of these inhibitors challenged the standard protease assay leading to a new screening platform implemented and validated. Reconnaissance studies from this new screening platform delineated an unprecedented structural flexibility associated with BoNT/A’s enzyme pocket, which can be induced by a small molecule for enhanced allosteric target inhibition of the protease. The culmination of these findings offers previously unrealized opportunities for neutralizing the BoNT/A protease and thus future <i>in vivo</i> applications.