eSrtA(2A-9) pET29b
(Plasmid #75145)

• Purpose: reprogrammed sortase A variant that recognizes alternative substrate LAXTG with high activity and specificity
• Depositing Lab: David Liu
• Publication: Dorr et al Proc Natl Acad Sci U S A. 2014 Sep 16;111(37):13343-8. doi: 10.1073/pnas.1411179111. Epub 2014 Sep 3.

Backbone

• Vector backbone: pET29b
• Backbone manufacturer: Novagen
• Backbone size w/o insert (bp): 5370
• Vector type: Bacterial Expression

Growth in Bacteria

• Bacterial Resistance(s): Kanamycin, 50 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: High Copy

Gene/Insert

• Gene/Insert name: reprogrammed sortase A variant
• Alt name: eSrtA 2A-9
• Species: Staphylococcus aureus
• Insert Size (bp): 441
• Mutation: S102C A104H E105D K138P K152I N160K K162H T164N K173E I182V T196S compared to eSrtA. Deleted amino acids 1-59 (which do not affect catalytic activity).
• Promoter: T7
• Tag / Fusion Protein:
  • 6x His (C terminal on backbone)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: NdeI (not destroyed)
• 3′ cloning site: BamHI (not destroyed)
• 5′ sequencing primer: T7
• 3′ sequencing primer: T7-term

Resource Information

• Supplemental Documents:
  • Annotated GenBank file
• Articles Citing this Plasmid:
  • 4 References

Terms and Licenses

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

Depositor Comments

eSrtA(2A-9) has 51,000 fold change in substrate specificity with activity comparable to eSrtA and it prefers LAXTG substrate over wild-type LPXTG. See sortase A pentamutant (eSrtA) in pET29 for original eSrtA (Addgene plasmid# 75144).