Structural Characterization of Three Novel Hydroxamate-Based Zinc Chelating Inhibitors of the Clostridium botulinum Serotype A Neurotoxin Light Chain Metalloprotease Reveals a Compact Binding Site Resulting from 60/70 Loop Flexibility

• Published in: Biochemistry (Easton) 2011-05, Vol.50 (19), p.4019-4028
• Main Authors: Thompson, Aaron A, Jiao, Guan-Sheng, Kim, Seongjin, Thai, April, Cregar-Hernandez, Lynne, Margosiak, Stephen A, Johnson, Alan T, Han, Gye Won, O’Malley, Sean, Stevens, Raymond C
• Format: Article
• Language: English
• Subjects: Binding Sites; Botulinum Toxins, Type A - antagonists & inhibitors; Botulinum Toxins, Type A - chemistry; Botulinum Toxins, Type A - metabolism; Chelating Agents - chemistry; Chelating Agents - metabolism; Clostridium botulinum - enzymology; Crystallography, X-Ray; Hydroxamic Acids - chemistry; Hydroxamic Acids - metabolism; Ligands; Metalloproteases - antagonists & inhibitors; Metalloproteases - chemistry; Metalloproteases - metabolism; Neurotoxins - antagonists & inhibitors; Neurotoxins - chemistry; Neurotoxins - metabolism; Protein Binding; Protein Conformation; Protein Structure, Tertiary; Zinc - chemistry; Zinc - metabolism
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/bi2001483

Neurotoxins synthesized by Clostridium botulinum bacteria (BoNT), the etiological agent of human botulism, are extremely toxic proteins making them high-risk agents for bioterrorism. Small molecule inhibitor development has been focused on the light chain zinc-dependent metalloprotease domain of the neurotoxin, an effort that has been hampered by its relatively flexible active site. Developed in concert with structure−activity relationship studies, the X-ray crystal structures of the complex of BoNT serotype A light chain (BoNT/A LC) with three different micromolar-potency hydroxamate-based inhibitors are reported here. Comparison with an unliganded BoNT/A LC structure reveals significant changes in the active site as a result of binding by the unique inhibitor scaffolds. The 60/70 loop at the opening of the active site pocket undergoes the largest conformational change, presumably through an induced-fit mechanism, resulting in the most compact catalytic pocket observed in all known BoNT/A LC structures.