Literature summary for 3.4.24.69 extracted from

Silvaggi, N.R.; Wilson, D.; Tzipori, S.; Allen, K.N.
Catalytic features of the botulinum neurotoxin A light chain revealed by high resolution structure of an inhibitory peptide complex (2008), Biochemistry, 47, 5736-5745.

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Application

Application	Comment	Organism
drug development	because of the potential for use of the toxin in bioterrorism and the increasingly widespread application of the toxin in the medical field, there is interest in the development of small-molecule inhibitors of the metalloprotease	Clostridium botulinum

Crystallization (Commentary)

Crystallization (Comment)	Organism
complex between the botulinum neurotoxin A light chain and the inhibitory peptide N-Ac-CRATKML, X-ray diffraction structure determination and analysis at 1.4 A resolution, and in a second approach unliganded enzyme light chain with and without the Zn2+ cofactor bound, X-ray diffraction structure determination and anaylsis at 1.25 A and 1.20 A resolution, respectively, 6-8 mg/ml purified BoNT/ALC, residues 1-425, in 50 mM NaPO4, pH 6.0, and 2 mM EDTA, hanging drop vapor diffusion method, mixing of protein solution with reservoir solution containing 20% PEG 3,350, 0.2 M diammonium tartrate, pH 6.6, and equilibration against 0.5 ml reservoir solution, the crystals are soaked prior to cryo-cooling in the crystallization solution plus 10 mM Zn(NO3)2 for 4.5 h or 5 mM Zn(NO3)2 and 2 mM N-Ac-CRATKML for 23 h, respectively, modeling	Clostridium botulinum

Crystallization (Comment)

Organism

complex between the botulinum neurotoxin A light chain and the inhibitory peptide N-Ac-CRATKML, X-ray diffraction structure determination and analysis at 1.4 A resolution, and in a second approach unliganded enzyme light chain with and without the Zn2+ cofactor bound, X-ray diffraction structure determination and anaylsis at 1.25 A and 1.20 A resolution, respectively, 6-8 mg/ml purified BoNT/ALC, residues 1-425, in 50 mM NaPO4, pH 6.0, and 2 mM EDTA, hanging drop vapor diffusion method, mixing of protein solution with reservoir solution containing 20% PEG 3,350, 0.2 M diammonium tartrate, pH 6.6, and equilibration against 0.5 ml reservoir solution, the crystals are soaked prior to cryo-cooling in the crystallization solution plus 10 mM Zn(NO3)2 for 4.5 h or 5 mM Zn(NO3)2 and 2 mM N-Ac-CRATKML for 23 h, respectively, modeling

Clostridium botulinum

Inhibitors

Inhibitors	Comment	Organism	Structure
N-Ac-CRATKML	an inhibitory peptide, structure of the serotype A toxin light chain with an inhibitory peptide bound at the catalytic Zn(II) ion, the peptide is bound with the Cys Sgamma atom coordinating the metal ion, overview	Clostridium botulinum

Localization

Localization	Comment	Organism	GeneOntology No.	Textmining

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Zn2+	BoNT/A-LC is a Zn(II)-dependent metalloprotease, the Zn2+ ion plays a purely catalytic role	Clostridium botulinum

Molecular Weight [Da]

Molecular Weight [Da]	Molecular Weight Maximum [Da]	Comment	Organism
50000	-	1 * 50000, light chain, + 1 * 100000, heavy chain, linked by a disulfide bond	Clostridium botulinum
100000	-	1 * 50000, light chain, + 1 * 100000, heavy chain, linked by a disulfide bond	Clostridium botulinum
150000	-	holotoxin	Clostridium botulinum

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
SNAP-25 + H2O	Clostridium botulinum	BoNT/A-LC is a Zn(II)-dependent metalloprotease that blocks the release of acetylcholine at the neuromuscular junction by cleaving SNAP-25, one of the SNARE proteins required for exocytosis	?	-	?

Organism

Organism	UniProt	Comment	Textmining
Clostridium botulinum	P10845	-	-

Posttranslational Modification

Posttranslational Modification	Comment	Organism
proteolytic modification	BoNTs are produced as a single 150 kDa polypeptide chain that is subsequently cleaved by endogenous proteases to give the dichain holotoxin	Clostridium botulinum

Reaction

Reaction	Comment	Organism	Reaction ID
Limited hydrolysis of proteins of the neuroexocytosis apparatus, synaptobrevin (also known as neuronal vesicle-associated membrane protein, VAMP), synaptosome-associated protein of 25 kDa (SNAP25) or syntaxin. No detected action on small molecule substrates	reaction mechanism	Clostridium botulinum	a

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
SNAP-25 + H2O	BoNT/A-LC is a Zn(II)-dependent metalloprotease that blocks the release of acetylcholine at the neuromuscular junction by cleaving SNAP-25, one of the SNARE proteins required for exocytosis	Clostridium botulinum	?	-	?
SNAP-25 + H2O	i.e. synaptosomal associated protein of 25 kDa	Clostridium botulinum	?	-	?

Subunits

Subunits	Comment	Organism
dimer	1 * 50000, light chain, + 1 * 100000, heavy chain, linked by a disulfide bond	Clostridium botulinum

Synonyms

Synonyms	Comment	Organism
BoNT/A-LC	-	Clostridium botulinum
botulinum neurotoxin A light chain	-	Clostridium botulinum