Serotype-Selective, Small-Molecule Inhibitors of the Zinc Endopeptidase of Botulinum Neurotoxin Serotype A [report]

Jewn G. Park, Peter C. Sill, Edward F. Makiyi, Alfonso T. Garcia-Sosa, Charles B. Millard, James J. Schmidt, Yuan-Ping Pang
2006 unpublished
Botulinum neurotoxin serotype A (BoNTA) is one of the most toxic substances known. Currently, there is no antidote to BoNTA. Small molecules identified from high-throughput screening reportedly inhibit the endopeptidase-the zinc-bound, catalytic domain of BoNTA-at a drug concentration of 20 lM. However, optimization of these inhibitors is hampered by challenges including the computational evaluation of the ability of a zinc ligand to compete for coordination with nearby residues in the active
more » ... ues in the active site of BoNTA. No improved inhibitor of the endopeptidase has been reported. This article reports the development of a serotype-selective, small-molecule inhibitor of BoNTA with a K i of 12 lM. This inhibitor was designed to coordinate the zinc ion embedded in the active site of the enzyme for affinity and to interact with a species-specific residue in the active site for selectivity. It is the most potent small-molecule inhibitor of BoNTA reported to date. The results suggest that multiple molecular dynamics simulations using the cationic dummy atom approach are useful to structure-based design of zinc protease inhibitors. Public reporting burden for the collection of information is estimated to average 1 hour per response, including the time for reviewing instructions, searching existing data sources, gathering and maintaining the data needed, and completing and reviewing the collection of information. Send comments regarding this burden estimate or any other aspect of this collection of information, including suggestions for reducing this burden, to Washington Headquarters Services, Directorate for Information Operations and Reports, 1215 Jefferson Davis Highway, Suite 1204, Arlington VA 22202-4302. Respondents should be aware that notwithstanding any other provision of law, no person shall be subject to a penalty for failing to comply with a collection of information if it does not display a currently valid OMB control number. 1. REPORT DATE no antidote to BoNTA. Small molecules identified from high-throughput screening reportedly inhibit the endopeptidase -the zinc-bound, catalytic domain of BoNTA -at a drug concentration of 20 µM. However, optimization of these inhibitors is hampered by challenges including the computational evaluation of the ability of a zinc ligand to compete for coordination with nearby residues in the active site of BoNTA. No improved inhibitor of the endopeptidase has been reported. This article reports the development of a serotype-specific small-molecule inhibitor of BoNTA with Ki of 12 µm. it suggests that multiple molecular dynamics simulations using the cationic dummy atom approach are useful to structure-based design of zinc protease inhibitors. 15 . SUBJECT TERMS countermeasures, antidotes, protease, structure-based drug desgin, Clostridium botulinum, neurotoxin 16. SECURITY CLASSIFICATION OF: 17. LIMITATION OF ABSTRACT SAR 18. NUMBER OF PAGES 14 19a. NAME OF RESPONSIBLE PERSON a. REPORT unclassified b. ABSTRACT unclassified c. THIS PAGE unclassified Standard Form 298 (Rev. 8-98) Prescribed by ANSI Std Z39-18 endopeptidase. The model suggests that the deep active site of the endopeptidase is filled with a rod-shaped heptapeptide. This structural feature requires that a molecule be at least 10 carbons long to be an effective inhibitor of the endopeptidase. This is consistent with the fact that small-molecule inhibitors reported to date are more than 10 carbons long. 11 The model also suggests that the interaction of Phe193 of the endopeptidase with a substrate or an inhibitor partly establishes specificity of a substrate or selectivity of inhibitors. Second, there are examples 12,13 to support the notion that the affinities of the small molecules binding at the active site of the endopeptidase could be greatly improved by these moleculesÕ coordination to the zinc ion at the active site. The presence of a zinc ion in the active site of the endopeptidase permits the design of small molecules with affinities that are high enough for these molecules to compete with protein substrates for binding to the endopeptidase. The affinities of the small molecules binding at the heavy-chain domain are, however, difficult to improve to the extent that these molecules can compete with neuronal cells for binding to the heavy-chain domain. This is because the heavy-chain domain lacks a deep pocket at the interface of its complex. 5,9
doi:10.21236/ada443596 fatcat:5h3rvccegnhpxpugdtprfsdw74