Part:BBa_K3746012
T7-LacO-NSP4-FDRA-6His
Description
This is a part design for the Phase I project (2021 iGEM) of Team Hong Kong JSS.
The IPTG inducible secretory expression construct of F420H2-dependent reductase (FDR)-A from Mycobacterium smegmatis (BBa_K3746003). It is the secretory version of expression construct (BBa_K3746010).
The coding sequence of tvLac is driven by T7 promoter (BBa_I719005), a Lac Opertator (BBa_K3076802), strong RBS (BBa_K3076801), a signal peptide for protein secretion (NSP4 (BBa_K3746004)) and a T7 terminator (BBa_K395601).
The FDR-A protein is tagged with a C-terminal 6-His Tag (CATCATCATCATCATCAT) for protein purification.
The NSP4 is a newly discovered signal peptide used in E. coli for protein secretion by signal recognition particle (SRP) pathway. This signal peptide was developed by Han, S.K., et al. (2017) based on the sequence of DsbAss signal peptide. They reported that this Novel Signal Peptide 4 has about 4 times higher secretory efficiency than the conventional DsbAss signal peptide. [1]
Reference
[1] Han, S. J., Machhi, S., Berge, M., Xi, G., Linke, T., & Schoner, R. (2017). Novel signal peptides improve the secretion of recombinant Staphylococcus aureus alpha toxinH35L in Escherichia coli. AMB Express, 7(1). https://doi.org/10.1186/s13568-017-0394-1
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 119
Illegal AgeI site found at 192 - 1000COMPATIBLE WITH RFC[1000]
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