Abstract
Clostridium botulinum neurotoxin is the most poisonous substance known to humans. It is a potential biowarfare threat and a public health hazard. The only therapeutics available is antibody treatment which will not be effective for post-exposure therapy. There are no drugs available for post-intoxication treatment. Accordingly, it is imperative to develop effective drugs to counter botulism. Available structural information on botulinum neurotoxins both alone and in complex with their substrates offers an efficient method for designing structure-based drugs to treat botulism.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Abbreviations
- BoNT:
-
Botulinum neurotoxin
- SNAP-25:
-
Synaptosomal-associated protein of 25Â kDa
- VAMP:
-
Vesicle-associated membrane protein
- SNARE:
-
Soluble N-ethylmaleimide-sensitive factor attachment protein receptor
- HC:
-
Heavy chain
- LC:
-
Light chain
References
Adler M, Nicholson JD, Hackley BE (1998) Efficacy of a novel metalloprotease inhibitor on botulinum neurotoxin B activity. FEBS Lett 429:234–238
Adler M, Nicholson JD, Starks DF, Kane CT, Cornille F, Hackley BE (1999) Evaluation of phosphoramidon and three synthetic phosphonates for inhibition of botulinum neurotoxin B catalytic activity. J Appl Toxicol 19:S5–S11
Adler MJ, Jamieson AG, Hamilton AD (2011) Hydrogen-bonded synthetic mimics of protein secondary structure as disruptors of protein–protein interactions. Curr Top Microbiol Immunol 348:1–23
Agarwal R, Binz T, Swaminathan S (2005a) Analysis of active site residues of botulinum neurotoxin E by mutational, functional and structural studies: Glu335Gln is an apoenzyme. Biochemistry 44:8291–8302
Agarwal R, Binz T, Swaminathan S (2005b) Structural analysis of botulinum neurotoxin serotype f light chain: implications on substrate binding and inhibitor design. Biochemistry 44:11758–11765
Agarwal R, Eswaramoorthy S, Kumaran D, Binz T, Swaminathan S (2004) Structural analysis of botulinum neurotoxin type E catalytic domain and its mutant Glu212->Gln reveals the pivotal role of the Glu212 carboxylate in the catalytic pathway. Biochemistry 43:6637–6644
Agarwal R, Schmidt JJ, Stafford RG, Swaminathan S (2009) Mode of VAMP substrate recognition and inhibition of Clostridium botulinum neurotoxin F. Nat Struct Mol Biol 16:789–794
Agarwal R, Swaminathan S (2008) SNAP-25 substrate peptide (residues 180–183) binds to but bypasses cleavage by catalytically active Clostridium botulinum neurotoxin E. J Biol Chem 283:25944–25951
Anne C, Turcaud S, Blommaert AG, Darchen F, Johnson EA, Roques BP (2005) Partial protection against Botulinum B neurotoxin-induced blocking of exocytosis by a potent inhibitor of its metallopeptidase activity. Chem Bio Chem 8:1375–1380
Anne C, Turcaud S, Quancard J, Teffo F, Meudal H, Fournie-Zaluski M-C, Roques BP (2003) Development of potent inhibitors of botulinum neurotoxin type B. J Med Chem 46:4648–4656
Arndt JW, Chai Q, Christian T, Stevens RC (2006) Structure of botulinum neurotoxin type D light chain at 1.65 A resolution: repercussions for VAMP-2 substrate specificity. Biochem Cell Biol 45:3255–3262
Arndt JW, Yu W, Bi F, Stevens RC (2005) Crystal structure of botulinum neurotoxin type G light chain: serotype divergence in substrate recognition. Biochemistry 44:9574–9580
Binz T, Bade S, Rummel A, Kollewe A, Alves J (2002) Arg362 and Tyr365 of the botulinum neurotoxin type A light chain are involved in transition state stabilization. Biochemistry 41:1717–1723
Boldt GE, Eubanks LM, Janda KD (2006a) Identification of a botulinum neurotoxin A protease inhibitor displaying efficacy in a cellular model. Chem Commun 7:3063–3065
Boldt GE, Kennedy JP, Hixon MS, McAllister LA, Barbieri JT, Tzipori S, Janda KD (2006b) Synthesis, characterization and development of a high-throughput methodology for the discovery of botulinum neurotoxin A inhibitors. J Comb Chem 8:513–521
Breidenbach MA, Brunger A (2004) Substrate recognition strategy for botulinum neurotoxin serotype A. Nature 432:925–929
Brewer M, Oost T, Sukonpan C, Pereckas M, Rich DH (2002) Sequencing hydroxyethylamine-containing peptides via Edman degradation. Org Lett 4:3469–3472
Brunger AT, Breidenbach MA, Jin R, Fischer A, Santos JS, Montal M (2007) Botulinum neurotoxin heavy chain belt as an intramolecular chaperone for the light chain. PLoS Pathog 3:1191–1194
Burnett JC, Ruthel G, Stegmann CM, Panchal RG, Nguyen TL, Hermone AR, Stafford RG, Lane DJ, Kenny TA, McGrath CF, Wipf P, Stahl AM, Schmidt JJ, Gussio R, Brunger AT, Bavari S (2007) Inhibition of metalloprotease botulinum serotype A from a pseudo-peptide binding mode to a small molecule that is active in primary neurons. J Biol Chem 282:5004–5014
Capkova K, Hixon MS, McAllister LA, Janda KD (2008) Toward the discovery of potent inhibitors of botulinum neurotoxin A: development of a robust LC MS based assay operational from low to subnanomolar enzyme concentrations. Chem Commun (Camb) 30:3525–3527
Capkova K, Hixon MS, Pellett S, Barbieri JT, Johnson EA, Janda KD (2010) Benzylidene cyclopentenediones: First irreversible inhibitors against botulinum neurotoxin A’s zinc endopeptidase. Bioorg Med Chem Lett 20:206–208
Capkova K, Salzameda NT, Janda KD (2009) Investigations into small molecule non-peptidic inhibitors of the botulinum neurotoxins. Toxicon 54:575–582
Capkova K, Yoneda Y, Dickerson TJ, Janda KD (2007) Synthesis and structure-activity relationships of second-generation hydroxamate botulinum neurotoxin A protease inhibitors. Bioorg Med Chem Lett 17:6463–6466
Chai Q, Arndt JW, Dong M, Tepp WH, Johnson EA, Chapmann ER, Stevens RC (2006) Structural basis of cell surface receptor recognition by botulinum neurotoxin B. Nature (London), New Biology 444:1019–1020
Chen S, Barbieri JT (2006) Unique substrate recognition by botulinum neurotoxins serotypes A and E. J Biol Chem 281:10906–10911
Chen S, Barbieri JT (2007) Multiple pocket recognition of SNAP25 by botulinum neurotoxin serotype E. J Biol Chem 282:25540–25547
Fischer A, Montal M (2007) Crucial role of the disulfide bridge between botulinum neurotoxin light and heavy chains in protease translocation across membranes. J Biol Chem 282:29604–29611
Fischer A, Mushrush DJ, Lacy DB, Montal M (2008) Botulinum neurotoxin devoid of receptor binding domain translocates active protease. PLoS Pathog 4:e1000245
Galloux M, Vitrac H, Montagner C, Raffestin S, Popoff MR, Chenal A, Forge V, Gillet D (2008) Membrane Interaction of botulinum neurotoxin A translocation (T) domain. The belt region is a regulatory loop for membrane interaction. J Biol Chem 283:27668–27676
Hale M, Oyler G, Swaminathan S, Ahmed SA (2010) Basic tetrapeptides as potent intracellular inhibitors of type A botulinum neurotoxin protease activity. J Biol Chem 286:1802–1811
Hermone AR, Burnett JC, Nuss JE, Tressler LE, Nguyen TL, Solaja BA, Vennerstorm JL, Schmidt JJ, Wipf P, Bavari S, Gussio R (2008) Three-Dimensional Database Mining Identifies a Unique Chemotype that Unites Structurally Diverse Botulinum Neurotoxin Serotype A Inhibitors in a Three-Zone Pharmacophore. Chem Med Chem 3:1905–1912
Jin R, Rummel A, Binz T, Brunger AT (2006) Botulinum neurotoxin B recognizes its protein receptor with high affinity and specificity. Nature (London) New Biology 444:1092–1095
Jin R, Sikorra S, Stegmann CM, Pich A, Binz T, Brunger AT (2007) Structural and biochemical studies of botulinum neurotoxin serotype C1 light chain protease: implications for dual substrate specificity. Biochemistry 46:10685–10693
Koriazova L, Montal M (2003) Translocation of botulinum neurotoxin light chain protease through the heavy chain channel. Nat Struct Biol 10:13–18
Kumar G, Agarwal R, Swaminathan S (2012a) Discovery of a fluorene class of compounds as inhibitors of botulinum neurotoxin serotype E by virtual screening. Chem Commun (Camb) 48:2412–2414
Kumar G, Kumaran D, Ahmed SA, Swaminathan S (2012b) Peptide inhibitors of botulinum neurotoxin serotype A: design, inhibition, co crystal structures, structure-activity relationship and pharmacophore modeling. Acta Crystallogr D Biol Crystallogr 68:511–520
Kumaran D, Eswaramoorthy S, Furey W, Navaza J, Sax M, Swaminathan S (2009) Domain organization in Clostridium botulinum neurotoxin type E is unique: Its implication in faster translocation. J Mol Biol 386:233–245
Kumaran D, Rawat R, Ahmed SA, Swaminathan S (2008a) Substrate binding mode and its implication on drug design for botulinum neurotoxin A. PLoS Pathog 4:e1000165
Kumaran D, Rawat R, Ludivico ML, Ahmed SA, Swaminathan S (2008b) Structure and substrate based inhibitor design for clostridium botulinum neurotoxin serotype A. J Biol Chem 283:18883–18891
Lacy DB, Tepp W, Cohen AC, DasGupta BR, Stevens RC (1998) Crystal structure of botulinum neurotoxin type A and implications for toxicity. Nat Struct Biol 5:898–902
Li L, Binz T, Niemann H, Singh BR (2000) Probing the mechanistic role of glutamate residues in the zinc-binding motif of type A botulinum neurotoxin light chain. Biochemistry 39:2399–2405
Li Y, Foran P, Fairweather NF, Paiva Ad, Weller U, Dougan G, Dolly JO (1994) A single mutation in the recombinant light chain of tetanus toxin abolishes its proteolytic activity and removes the toxicity seen after reconstitution with native heavy chain. Biochemistry 33:7014–7020
Martin L, Cornille F, Turcaud S, Meudal H, Roques BP, Fourine-Zaluski MC (1999) Metallopeptidase inhibitors of tetanus toxin: a combinatorial approach. J Med Chem 42:515–525
Masuyer G, Thiyagarajan N, James PL, Marks PHH, Chaddock J, Acharya KR (2009) Crystal structure of a catalytically active, non-toxic endopeptidase derivative of Clostridium botulinum toxin A. Biochem Biophys Res Commun 381:50–53
Matthews BW (1988) Structural basis of the action of thermolysin and related zinc peptides. Acc Chem Res 21:333–340
Montal M (2009) Translocation of botulinum neurotoxin light chain protease by the heavy chain protein-conducting channel. Toxicon 54:565–569
Montecucco C, Papini E, Schiavo G (1994) Bacterial protein toxins penetrate cells via a four-step mechanism. FEBS Lett 346:92–98
Oost T, Sukonpan C, Brewer M, Goodnough M, Tepp W, Johnson EA, Rich DH (2003) Design and synthesis of substrate-based inhibitors of botulinum neurotoxin type B metalloprotease. Biopolymers 71:602–619
Pang YP, Davis J, Wang S, Park JG, Nambiar MP, Schmidt JJ, Millard CB (2010a) Small molecules showing significant protection of mice against botulinum neurotoxin serotype A. PLoS ONE 5:e10129
Pang YP, Vummenthala A, Mishra RK, Park JG, Wang S, Davis J, Millard CB, Schmidt JJ (2009) Potent new small-molecule inhibitor of botulinum neurotoxin serotype A endopeptidase developed by synthesis-based computer-aided molecular design. PLoS ONE 4:e7730
Pang YP, Vummenthala A, Mishra RK, Park JG, Wang S, Davis J, Millard CB, Schmidt JJ (2010b) Potent new small-molecule inhibitor of botulinum neurotoxin serotype A endopeptidase developed by synthesis-based computer-aided molecular design. PLoS ONE 4:e7730
Rummel A, Eichner T, Weil T, Karnath T, Gutcaits A, Mahrhold S, Sandhoff K, Proia RL, Acharya KR, Bigalke H, Binz T (2007) Identification of the protein receptor binding site of botulinum neurotoxins B and G proves the double-receptor concept. Proc Natl Acad Sci USA 104:359–364
Salzameda NT, Eubanks ME, Zakhari JS, Tsuchikama K, DeNunzio NJ, Allen KA, Hixon SS, Janda KD (2011) A cross-over inhibitor of the botulinum neurotoxin light chain B: a natural product implicating an exosite mechanism of action. Chem Commun 47:1713–1715
Schiavo G, Matteoli M, Montecucco C (2000) Neurotoxins affecting neuroexocytosis. Physiol Rev 80:717–766
Schmidt JJ, Stafford RG (2002) A high-affinity competitive inhibitor of type A botulinum neurotoxin protease activity. FEBS Lett 532:423–426
Schmidt JJ, Stafford RG (2005) Botulinum neurotoxin serotype F: identification of substrate recognition requirements and development of inhibitors with low nanomolar affinity. Biochemistry 44:4067–4073
Schmidt JJ, Stafford RG, Bostian KA (1998) Type A botulinum neurotoxin proteolytic activity: development of competitive inhibitors and implications for substrate specificity at the S1’ binding subsite. FEBS Lett 435:61–64
Sikorra S, Henke T, Galli T, Binz T (2008) Substrate recognition mechanism of VAMP/synaptobrevin-cleaving clostridial neurotoxins. J Biol Chem 283:21145–21152
Silhár P, Capková K, Salzameda NT, Barbieri JT, Hixon MS, Janda KD (2010) Botulinum neurotoxin A protease: discovery of natural product exosite inhibitors. J Am Chem Soc 132:2868–2869
Silvaggi NR, Wilson D, Tzipori S, Allen KN (2008) Catalytic features of the botulinum neurotoxin a light chain revealed by high resolution structure of an inhibitory peptide complex. Biochemistry 47:5736–5745
Stenmark P, Dupuy J, Imamura A, Kiso M, Stevens RC (2008) Crystal structure of botulinum neurotoxin type A in complex with the cell surface co-receptor GT1b-insight into the toxin-neuron interaction. PLoS Pathog 4:e1000129
Sukonpan C, Oost T, Goodnough M, Tepp W, Johnson EA, Rich DH (2004) Synthesis of substrates and inhibitors of botulinum neurotoxin type A metalloprotease. J Pept Res 63:181–193
Swaminathan S, Eswaramoorthy S (2000) Structural analysis of the catalytic and binding sites of Clostridium botulinum neurotoxin B. Nat Struct Biol 7:693–699
Swaminathan S, Eswaramoorthy S, Kumaran D (2004) Structure and enzymatic activity of botulinum neurotoxins. Movement Disord 19(8):S17–S22
Thompson AA, Jiao GS, Kim S, Thai A, Cregar-Hernandez L, Margosiak SA, Johnson AT, Han GW, O’Malley S, Stevens RC (2011) 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. Biochemistry 50:4019–4028
Vaidyanathan VV, Yoshino K, Jahnz M, Dorries C, Bade S, Nauenburg S, Niemann H, Binz T (1999) Proteolysis of SNAP-25 isoforms by botulinum neurotoxin types A, C, and E: Domains and amino acid residues controlling the formation of enzyme-substrate complexes and cleavage. J Neurochem 72:327–337
Zuniga JE, Hammill JT, Drory O, Nuss JE, Burnett JC, Gussio R, Wipf P, Bavari S, Brunger AT (2010) Iterative structure-based peptide-like inhibitor design against the botulinum neurotoxin serotype A. PLoS ONE 5:e11378
Zuniga JE, Schmidt JJ, Fenn T, Burnett JC, Arac D, Gussio R, Stafford RG, Badie SS, Bavari S, Brunger AT (2008) A potent peptidomimetic inhibitor of botulinum neurotoxin serotype A has a very different conformation than SNAP-25 substrate. Structure (Camb) 16:1588–1597
Acknowledgments
The author is thankful to his colleagues and collaborators for contributing to this research. Special thanks are due to Drs. S. Eswaramoorthy, D. Kumaran, R. Agarwal, S.A. Ahmed, and G. Kumar. Research was supported by an award from DTRA BO742081 under DOE prime contract No. DEAC02-98CH10886 with Brookhaven National Laboratory.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2012 Springer-Verlag Berlin Heidelberg
About this chapter
Cite this chapter
Swaminathan, S. (2012). Structure-Based Drug Discovery for Botulinum Neurotoxins. In: Rummel, A., Binz, T. (eds) Botulinum Neurotoxins. Current Topics in Microbiology and Immunology, vol 364. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-3-642-33570-9_10
Download citation
DOI: https://doi.org/10.1007/978-3-642-33570-9_10
Published:
Publisher Name: Springer, Berlin, Heidelberg
Print ISBN: 978-3-642-33569-3
Online ISBN: 978-3-642-33570-9
eBook Packages: Biomedical and Life SciencesBiomedical and Life Sciences (R0)