SpyTag/SpyCatcher Protein Coupling Reagents

The SpyTag/SpyCatcher system is a convenient protein coupling tool for irreversible peptide-protein ligation. It is ideal for binding, labeling, immobilization and creating new kinds of protein architectures.

Highlights:

  • SpyTag forms a spontaneous amide bond upon binding its genetically encoded partner SpyCatcher
  • Highly compatible - SpyTag reacts with SpyCatcher under a wide range of conditions
  • Extremely stable reaction product - SpyCatcher/SpyTag complex is stable to boiling in SDS
  • Cys-SpyCatcher version contains N-terminal cysteine for precise labeling with dye or precise attachment to surfaces or beads

Peptide interaction with proteins is usually weak. SpyTag is a genetically encoded peptide (sequence AHIVMVDAYKPTK) that forms a spontaneous amide bond upon binding its genetically encoded partner SpyCatcher. SpyTag reacts with SpyCatcher under a wide range of conditions and the after reaction the product is stable to boiling in SDS. Cys-SpyCatcher has a unique N-terminal cysteine for precise labeling with dye or precise attachment to surfaces or beads.

From the laboratory of Mark Howarth, PhD, University of Oxford.

The Investigator's Annexe Part of The Investigator's Annexe program.

Catalog Number Product Size AVAILABILITY Price Qty
EOX003
SpyCatcher Protein, 1mg
1mg (5.0mg/mL) Currently unavailable
Price: $154.00
EOX004
Cys-SpyCatcher Protein, 1mg
1mg (6.9mg/mL) Currently unavailable
Price: $154.00
EOX005
SUMO-SpyTag Protein, 1mg
1mg (9.9mg/mL) Currently unavailable
Price: $154.00
Specifications
Product Type: Protein
Name: SpyTag/SpyCatcher Protein Coupling Reagents
Accession ID: JQ478411.1 (SpyCatcher)
Fusion Tag(s): N-terminal 6xHis tag for each
Molecular Weight: 15,240.5 Da (SpyCatcher) 15,180.5 Da (Cys-SpyCatcher); 13,973.6 Da (SUMO-SpyTag)
Amino Acid Sequence: SpyCatcher Protein: SYYHHHHHHDYDIPTTENLYFQGAMVDTLSGLSSEQGQSGDMTIEEDSATHIKFSKRDEDGKELAGATMELRDSSGKTISTWISDGQVKDFYLYPGKYTFVETAAPDGYEVATAITFTVNEQGQVTVNGKATKGDAHI
Cys-SpyCatcher Protein: SYYHHHHHHDCDIPTTENLYFQGAMVDTLSGLSSEQGQSGDMTIEEDSATHIKFSKRDEDGKELAGATMELRDSSGKTISTWISDGQVKDFYLYPGKYTFVETAAPDGYEVATAITFTVNEQGQVTVNGKATKGDAHI

SUMO-SpyTag: GSSHHHHHHGSDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPLRRLMEAFAKRQGKEMDSLRFLYDGIRIQADQTPEDLDMEDNDIIEAHREQIGGGAHIVMVDAYKPTKGY

Buffer: Phosphate buffered saline (PBS) pH 7.5, 0.5mM EDTA
Source: Recombinant expression in E. coli
Purity: >95%. Purified from Ni-NTA affinity chromatography and then ion-exchange chromatography.
Concentration: 5.0mg/mL (SpyCatcher), 6.9mg/mL (Cys-SpyCatcher), 9.9mg/mL (SUMO-SpyTag)
Storage: -80C, avoid multiple freeze-thaw cycles
Shipped: Dry ice
Provider
From the laboratory of Mark Howarth, PhD, University of Oxford.
Comments

SpyCatcher reacts with SpyTag in the pH range 5-8 and is relatively insensitive to the buffer composition, as long as SDS or denaturants like urea or guanidinium are not used.

SpyCatcher: SDS-PAGE shows one higher band from gluconylation, which does not affect reactivity with SpyTag.

  • Theoretical pI: 4.85
  • Extinction coefficient at 280 nm (predicted by ProtParam): 17,420 M-1 cm-1

Cys-SpyCatcher: SDS-PAGE shows a 2nd band at slightly higher MW related to glucosylation, but this does not affect reactivity. As expected for exposed cysteines, Cys-SpyCatcher will eventually form a disulfide upon storage. This disulfide-bonded form will not be reactive with maleimide or other thiol-reactive reagents. To test the extent of disulfide bond formation, perform SDS-PAGE with Coomassie staining with and without DTT.

  • Theoretical pI: 4.85
  • Extinction coefficient at 280 nm (predicted by ProtParam): 15,930 M-1 cm-1

SUMO-SpyTag:

  • Theoretical pI: 5.97
  • Extinction coefficient at 280 nm (predicted by ProtParam): 4,470 M-1 cm-1

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