BoNT/A-LC
(Plasmid #31602)

• Depositing Lab: Axel Brunger
• Publication: Zuniga et al Structure. 2008 Oct 8. 16(10):1588-97.

Backbone

• Vector backbone: pBN3
• Vector type: Bacterial Expression

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: Unknown

Gene/Insert

• Gene/Insert name: BoNT/A LC
• Alt name: BoNT/A
• Tag / Fusion Protein:
  • His6 (C terminal on insert)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: None (unknown if destroyed)
• 3′ cloning site: None (unknown if destroyed)
• 5′ sequencing primer: n/a

Resource Information

• Article Citing this Plasmid:
  • 1 Reference

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
• Industry Terms:
  • Not Available to Industry

Depositor Comments

The author states that although the construct is from a toxic protein, it only contains one third of the biologically active protein,i.e., it is inactive and harmless per se, and it does not require BL4 level for its expression/manipulation.

Toxic BoNT/A is made of two 'chains' (i.e. polypeptides) linked to each other by one disulfide bond. The light chain (LC) is 50KDa long, whereas the heavy chain (HC) is 100 KDa. The HC is necessary for the toxin to target and enter motorneurons. This construct is the LC of BoNTA, and it completely lacks the HC, so it is unable to intoxicate neurons.

Protein sequence of BoNT/A fragment present in this plasmid (corresponds to YP_001386738.1)
MSGLEVSFEELRTFGGHDAKFIDSLQENEFRLYYYNKFKDIASTLNKAKSIVGTTASLQYMKNVFKEKYLLSEDTSGKFSVDKLKFDKLYKMLTEIYTEDNFVKFFKVLNRKTYLNFDKAVFKINIVPKVNYTIYDGFNLRNTNLAANFNGQNTEINNMNFTKLKNFT

Nucleotide sequence of BoNT/A fragment present in this plasmid based on Addgene's Sanger sequencing results:
ATGAGTGGGTTAGAAGTAAGCTTTGAGGAACTTAGAACATTTGGGGGACATGATGCAAAGTTTATAGATAGTTTACAGGAAAACGAATTTCGTCTATATTATTATAATAAGTTTAAAGATATAGCAAGTACACTTAATAAAGCTAAATCAATAGTAGGTACTACTGCTTCATTACAGTATATGAAAAATGTTTTTAAAGAGAAATATCTCCTATCTGAAGATACATCTGGAAAATTTTCGGTAGATAAATTAAAATTTGATAAGTTATACAAAATGTTAACAGAGATTTACACAGAGGATAATTTTGTTAAGTTTTTTAAAGTACTTAACAGAAAAACATATTTGAATTTTGATAAAGCCGTATTTAAGATAAATATAGTACCTAAGGTAAATTACACAATATATGATGGATTTAATTTAAGAAATACAAATTTAGCAGCAAACTTTAATGGTCAAAATACAGAAATTAATAATATGAATTTTACTAAACTAAAAAATTTTACT