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PDBsum entry 3c8b
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Hydrolase/hydrolase inhibitor
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PDB id
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3c8b
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References listed in PDB file
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Key reference
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Title
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Structure- And substrate-Based inhibitor design for clostridium botulinum neurotoxin serotype a.
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Authors
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D.Kumaran,
R.Rawat,
M.L.Ludivico,
S.A.Ahmed,
S.Swaminathan.
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Ref.
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J Biol Chem, 2008,
283,
18883-18891.
[DOI no: ]
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PubMed id
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Abstract
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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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Figure 4.
FIGURE 4. Schematic diagram of molecular interactions
between Balc424 and tetrapeptide inhibitors. The interactions
between Balc424 active site residues and inhibitor peptides are
shown. A, RRGC; B, RRGM; C, RRGL; and D, RRGI. Black, red, blue,
yellow, and green colored circles represent carbon, oxygen,
nitrogen, sulfur, and sodium atoms, respectively. For clarity,
zinc coordination and water molecules involved in the
interactions at the active site are not shown. The figure was
prepared with Ligplot (47).
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Figure 7.
FIGURE 7. Electrostatic potential surface representation of
active site cavity. Models of RRGC (green), RRGM (light green),
RRGL (coral), and RRGI (magenta) with sulfate ions are
superposed. Red and blue indicate negative (  -0.5 V) and positive (
 +0.5
V) electrostatic potential. The figure was prepared with CCP4MG
(50). The orientation is similar to Fig. 3A.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2008,
283,
18883-18891)
copyright 2008.
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