SUMO CRYGS 16-177 Q16C
(Plasmid #217673)

• Purpose: N-terminally truncated human gamma S crystallin residues 16-177 with Q16C mutation for native chemical ligation
• Depositing Lab: Larry David
• Publication: Completion of previous SUMO tagged human crystallin submission plus plasmids for native chemical ligation to produced gamma S crystallin (unpublished)

Backbone

• Vector backbone: pET28B+
• Backbone size w/o insert (bp): 5368
• Total vector size (bp): 5854
• Modifications to backbone: none
• 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: CRYGS
• Species: H. sapiens (human)
• Mutation: Glutamine 16 changed to Cysteine, and amino acids 1-15 removed from gamma S crystallin
• GenBank ID: CRYGS
• Entrez Gene: CRYGS (a.k.a. CRYG8, CTRCT20)
• Promoter: T7
• Tags / Fusion Proteins:
  • 6XHis (N terminal on backbone)
  • SUMO (N terminal on backbone)

Cloning Information

• Cloning method: Unknown
• 5′ sequencing primer: T7
• 3′ sequencing primer: T7 term

Resource Information

• A portion of this plasmid was derived from a plasmid made by: DNASU HsCD00404131

Terms and Licenses

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

Depositor Comments

This a N-terminally truncated gamma S crystallin missing its N-terminal 15 amino acid residues and containing a Q16C mutation at its N-terminus to act as a site to perform native chemical ligation. It also has a N-terminal SUMO tag with a 6X His tag with the amino acid sequence: GHHHHHHGSLQDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPLRRLMEAFAKRQGKEMDSLRFLYDGIRIQADQAPEDLDMEDNDIIEAHREQ IGG

It will express in a soluble state with the SUMO tag, but will precipitate once the SUMO tag is removed during ulp-1 cleavage. It has a propensity to form non-disulfide linked multimers once the SUMO tag is removed, so the ligation reaction must be performed immediately after ULP-1 treatment.