Part:BBa_K2273066

SpoIID signal peptide of B. subtilis lytic transglycosylase

The SpoIID signal peptide is part of the Signal Peptide Toolbox of [http://2017.igem.org/Team:TU_Dresden iGEM Team TU Dresden 2017 (EncaBcillus - It's a trap!)].

The signal peptide (amino acids 1 to 34) of Bacillus subtilis lytic transglycosylase (cell wall hydrolase involved in dissolution of the septal cell wall, Uniprot [http://www.uniprot.org/uniprot/P07372 P07372]) targets for protein secretion via the Sec-SRP secretion pathway (Brockmeier et al, 2006).

This part was generated in a modified version of RFC25, where a strong Shine Dalgarno Sequence (SD) is included, and has the following prefix and suffix:

Prefix with	EcoRI, NotI, XbaI and SD	GAATTCGCGGCCGCTTCTAGATAAGGAGGTCAAAA
Suffix with	AgeI, SpeI, NotI and PstI	ACCGGTTAATACTAGTAGCGGCCGCTGCAGA

Sites of restriction enzymes generating compatible overhangs are indicated by sharing one color. (EcoRI and PstI are marked in blue, NotI in green, XbaI and SpeI in red and AgeI in orange. Additionally, the Shine-Dalgarno sequence is marked in silver and the stop codon is underlined.)

This part is used in the 2017 TU Dresden iGEM project [http://2017.igem.org/Team:TU_Dresden EncaBcillus - It's a trap!] and part of the Signal Peptide Toolbox.

Signal Peptide Toolbox

The GRAM-positive model organism Bacillus subtilis is considered a perfect host for heterologous and recombinant protein secretion due to its extracellular chaperones, natural protein secretion capacity and well-studied genetics (van Dijl and Hecker, 2013). To increase protein production rates of such proteins, it is feasible to enhance the secretion efficiency. The easiest method to realise this, is to tag the protein of interest with a so-called signal peptide of B. subtilis' general protein secretion pathway Sec-SRP which dictates the secretion of proteins into the surrounding supernatant of the cell (Fu et al, 2007).

Though the Sec-SRP protein secretion pathway of B. subtilis contains more than 170 distinct signal peptides, every single SP varies in its secretion efficiency in dependency on the protein fused to it. To date, it is impossible to predict this dependency via studying the genetic code of the SPs and the protein of interest (Brockmeier et al, 2006).

To cope with this issue, the [http://2017.igem.org/Team:TU_Dresden iGEM team of TU Dresden 2017 (EncaBcillus - It's a trap!)] developed a novel shotgun approach high-throughput signal peptide screening vector system for targeted secretion in B. subtilis, or “Signal Peptide Toolbox” for short.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]