|RFC standard||RFC 25|
|Partner Tag||BBa_K2273015: mini. SpyCatcher|
|Original Biobrick Part||BBa_K1159201: SpyTag|
|His-tagged SpyTag||BBa_K2273016: SpyTag-His|
|Submitted by||TU Dresden|
SpyTag codon adapted for Bacillus subtilis
This is the codon optimized version of Biobrick BBa_K1159201 improved for ideal production in Bacillus subtilis. It codes for an oligopeptide forming an covalent isopeptide bond with a protein called SpyCatcher BBa_K1159200. There is also an shortened and codon optimized version of this protein called the Minimal SpyCatcher BBa_K2273015. The Spytag and the Minimal SpyCatcher can be fused as a tag to proteins of interest and will then mediate covalent bonding of these proteins.
The original part was used in E. coli but the SpyTag was thought to be implemented throughout TU_Dresdens project in B. subtilis. Therefore the sequence was codon adapted for this purposes. The sequence was proposed by researchersthat investigated the secretion of SpyTag/SpyCatcher system in B. subtilis.
The SpyTag was also submitted to the registry with an added C-terminal His-Tag to purify the protein constructs when SpyTag/SpyCatcher link together.
The 2017 team of TU_Dresden used the SpyTag together with its partner mini. SpyCatcher to investigate the potential of using B. subtilis secretion system for the production of self conjugation multi protein complexes. The his-tagged SpyTag was C-terminally fused to mCherry which functions as a reporter protein. The secretion was mediated through the N-terminal signal peptide of AmyE. The secretion of the functional mCherry fusion protein could be proven (Figure 1 and Figure 2). The functionality of the SpyTagr was proven through SDS-Page (Figure 3). After incubating supernatants containing mCherry-mini. SpyCatcher and mCherry-SpyTag the fusion product was detected, proving the covalent bonding of the Tag partners.
Sequence and Features
- 10COMPATIBLE WITH RFC
- 12COMPATIBLE WITH RFC
- 21COMPATIBLE WITH RFC
- 23COMPATIBLE WITH RFC
- 25COMPATIBLE WITH RFC
- 1000COMPATIBLE WITH RFC