Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.24.69 (botulinum neurotoxin)
 1,901 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The structure of botulinum neurotoxin type B (BoNT/B) is analyzed, and it is demonstrated that the carbonyl oxygen of the scissile bond comes close to the zinc ion to form a Michaelis complex. The hydrated carbonyl is activated by the nucleophilic water, which moves closer to Glu 230 to form hydrogen bonds to side-chain carboxylate. This process frees up the lone pair, which forms a bond with carbonyl carbon, corresponding to the tetrahedral transition state. The hydrated peptide oxygen is stabilized by a zinc ion and a water molecule close by. The proton from the nucleophile moves to NH of the scissile bond. The other proton is shuttled by Glu 230 to the NH2 group to make it NH3+ and allows it to leave. This mechanism is consistent with that proposed for thermolysin and BoNT/A. On the basis of these studies, we have shown that Tyr372 or Arg369 may not have any significant role in catalytic activity except for a secondary role such as stabilizing the transition state. Thus, the sulfate ion mimics the transition state of the scissile carbonyl carbon atom. However, the sulfate ion by itself does not inhibit the toxicity.
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PMID:Structure and enzymatic activity of botulinum neurotoxins. 1502 50

The seven antigenically distinct serotypes of Clostridium botulinum neurotoxins cleave specific soluble N-ethylmaleimide-sensitive factor attachment protein receptor complex proteins and block the release of neurotransmitters that cause flaccid paralysis and are considered potential bioweapons. Botulinum neurotoxin type A is the most potent among the clostridial neurotoxins, and to date there is no post-exposure therapeutic intervention available. To develop inhibitors leading to drug design, it is imperative that critical interactions between the enzyme and the substrate near the active site are known. Although enzyme-substrate interactions at exosites away from the active site are mapped in detail for botulinum neurotoxin type A, information about the active site interactions is lacking. Here, we present the crystal structures of botulinum neurotoxin type A catalytic domain in complex with four inhibitory substrate analog tetrapeptides, viz. RRGC, RRGL, RRGI, and RRGM at resolutions of 1.6-1.8 A. These structures show for the first time the interactions between the substrate and enzyme at the active site and delineate residues important for substrate stabilization and catalytic activity. We show that OH of Tyr(366) and NH(2) of Arg(363) are hydrogen-bonded to carbonyl oxygens of P1 and P1' of the substrate analog and position it for catalytic activity. Most importantly, the nucleophilic water is replaced by the amino group of the N-terminal residue of the tetrapeptide. Furthermore, the S1' site is formed by Phe(194), Thr(215), Thr(220), Asp(370), and Arg(363). The K(i) of the best inhibitory tetrapeptide is 157 nm.
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PMID:Structure- and substrate-based inhibitor design for Clostridium botulinum neurotoxin serotype A. 1843 12

Botulinum neurotoxins have a very high affinity and specificity for their target cells requiring two different co-receptors located on the neuronal cell surface. Different toxin serotypes have different protein receptors; yet, most share a common ganglioside co-receptor, GT1b. We determined the crystal structure of the botulinum neurotoxin serotype A binding domain (residues 873-1297) alone and in complex with a GT1b analog at 1.7 A and 1.6 A, respectively. The ganglioside GT1b forms several key hydrogen bonds to conserved residues and binds in a shallow groove lined by Tryptophan 1266. GT1b binding does not induce any large structural changes in the toxin; therefore, it is unlikely that allosteric effects play a major role in the dual receptor recognition. Together with the previously published structures of botulinum neurotoxin serotype B in complex with its protein co-receptor, we can now generate a detailed model of botulinum neurotoxin's interaction with the neuronal cell surface. The two branches of the GT1b polysaccharide, together with the protein receptor site, impose strict geometric constraints on the mode of interaction with the membrane surface and strongly support a model where one end of the 100 A long translocation domain helix bundle swing into contact with the membrane, initiating the membrane anchoring event.
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PMID:Crystal structure of botulinum neurotoxin type A in complex with the cell surface co-receptor GT1b-insight into the toxin-neuron interaction. 1870 64

The seven antigenically distinct serotypes of Clostridium botulinum neurotoxins, the causative agents of botulism, block the neurotransmitter release by specifically cleaving one of the three SNARE proteins and induce flaccid paralysis. The Centers for Disease Control and Prevention (CDC) has declared them as Category A biowarfare agents. The most potent among them, botulinum neurotoxin type A (BoNT/A), cleaves its substrate synaptosome-associated protein of 25 kDa (SNAP-25). An efficient drug for botulism can be developed only with the knowledge of interactions between the substrate and enzyme at the active site. Here, we report the crystal structures of the catalytic domain of BoNT/A with its uncleavable SNAP-25 peptide (197)QRATKM(202) and its variant (197)RRATKM(202) to 1.5 A and 1.6 A, respectively. This is the first time the structure of an uncleavable substrate bound to an active botulinum neurotoxin is reported and it has helped in unequivocally defining S1 to S5' sites. These substrate peptides make interactions with the enzyme predominantly by the residues from 160, 200, 250 and 370 loops. Most notably, the amino nitrogen and carbonyl oxygen of P1 residue (Gln197) chelate the zinc ion and replace the nucleophilic water. The P1'-Arg198, occupies the S1' site formed by Arg363, Thr220, Asp370, Thr215, Ile161, Phe163 and Phe194. The S2' subsite is formed by Arg363, Asn368 and Asp370, while S3' subsite is formed by Tyr251, Leu256, Val258, Tyr366, Phe369 and Asn388. P4'-Lys201 makes hydrogen bond with Gln162. P5'-Met202 binds in the hydrophobic pocket formed by the residues from the 250 and 200 loop. Knowledge of interactions between the enzyme and substrate peptide from these complex structures should form the basis for design of potent inhibitors for this neurotoxin.
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PMID:Substrate binding mode and its implication on drug design for botulinum neurotoxin A. 1881 39

The conformational property of oligosaccharide GT1B in aqueous environment was studied by molecular dynamics (MD) simulation using all-atom model. Based on the trajectory analysis, three prominent conformational models were proposed for GT1B. Direct and water-mediated hydrogen bonding interactions stabilize these structures. The molecular modeling and 15 ns MD simulation of the Botulinum Neuro Toxin/B (BoNT/B) - GT1B complex revealed that BoNT/B can accommodate the GT1B in the single binding mode. Least mobility was seen for oligo-GT1B in the binding pocket. The bound conformation of GT1B obtained from the MD simulation of the BoNT/B-GT1B complex bear a close conformational similarity with the crystal structure of BoNT/A-GT1B complex. The mobility noticed for Arg 1268 in the dynamics was accounted for its favorable interaction with terminal NeuNAc. The internal NeuNAc1 tends to form 10 hydrogen bonds with BoNT/B, hence specifying this particular site as a crucial space for the therapeutic design that can restrict the pathogenic activity of BoNT/B.
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PMID:Conformational analysis of GT1B ganglioside and its interaction with botulinum neurotoxin type B: a study by molecular modeling and molecular dynamics. 2274 33

The conformational property for the oligosaccharide structure of GD1A and GD1B in aqueous environment is studied by 10 ns Molecular Dynamics simulation using all atom model. Based on the trajectory analysis four conformational models are proposed for GD1A and one for GD1B. Direct and water mediated hydrogen bonding interactions plays a prominent role in stabilizing these conformational structures. The Molecular Modelling and 10 ns MD simulation of Botulinum Neuro Toxin/B-GD1A and BoNT/B-GD1B complex revealed that this toxin can interact with GD1A in the single binding mode and with GD1B in two binding modes. Least mobility is seen for GD1A in the binding pocket of BoNT/B. The GTSM comparison, pair interaction energy calculation, total energy calculation, MM/PBSA binding free energy calculation and RMSD predicts that GD1A is a better receptor for BoNT/B compared to GD1B. The internal NeuNAc1 tends to form more than 70% of hydrogen bonds with BoNT/B both in GD1A and GD1B, hence specifying this particular site as a crucial space for the therapeutic design that can restrict the pathogenic activity of BoNT/B.
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PMID:Molecular Modelling and Molecular Dynamics studies of GD1A, GD1B and their complexes with BoNT/B--perspectives in interaction and specificity. 2292 25