Recombinant Synaptobrevin-2 Protein Substrate for C. botulinum Types B, D, and F fused with GST tag was expressed in E. coli.
Tetanus toxin is a potent neurotoxin that is taken up by presynaptic terminals at the neuromuscular junction, travels retrogradely within motor neurons and is transported transsynaptically to spinal cord interneurons. Upon entering the motor neuron, tetanus toxin associates with lipid rafts, is incorporated into non-acidified vesicles, sorted in a Rab 5/7 dependent pathway and transported to the cell body. The tetanus toxin is trancytosed so that it enters upstream inhibitory neurons where it cleaves VAMP-2, inhibiting neurotransmitter release. Tetanus toxin inhibits neurotransmitter release with great potency and selectivity, cleaving a bond in vesicle-associated membrane protein-2 (VAMP-2) or synaptobrevin-2, thereby preventing the fusion of synaptic vesicles with the presynaptic membrane. Tetanus toxoid is currently used as a vaccine to produce immunity to tetanus, the disease caused by Clostridium tetani. Additionally, tetanus toxoid functions as a carrier of conjugated polysaccharides in the development of vaccines against encapsulated bacterial pathogens.
Each 100 ug vial of recombinant GST-Synaptobrevin-2, when reconstituted with 185 ul of water, contains 100 ug of protein in 20 mM HEPES, pH 7.4, 1.25% lactose.
The recombinant light chain from botulinum neurotoxin Type D (BoNT/D LC) (Product #630A) has been used to test the GST-Synaptobrevin-2 in an endopeptidase assay. Twenty nanomolar BoNT/D LC was incubated with 5 uM GST-Synaptobrevin-2 in 20 mM Tris, pH 8.0, containing 50 mM NaCI and 5 mM DTT at 37°C. In 1 hour, 95% of the GST-Synaptobrevin-2 was shown to be cleaved using SDS PAGE analysis.
GST-Synaptobrevin-2 is 331 amino acids in length. This product contains the GST affinity tag and 97 amino acids of the 116 amino acids of the Synaptobrevin-2 sequence. The final 19 amino acids of the native protein, containing the transmembrane domain, are not included in this preparation of Synaptobrevin-2.
331 amino acids
This product is supplied as a lyophilized powder which has been stoppered under vacuum. Store lyophilized vials at 2-8 centigrade. Once dissolved, aliquot and store the product at -20 centigrade. Refrain from multiple freeze/thaw cycles.
Good laboratory technique should be employed in the safe handling of this product. Wear appropriate laboratory attire including a lab coat, gloves and safety glasses. Nitrile gloves are recommended when handling lyophilized material. This product is intended for research purposes by qualified personnel only. It is not intended for use in humans or as a diagnostic agent.