Part:BBa_K4744777:Design
mostcomm-Strep-17-LCI
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
The sequence was included in a plasmid pET28a_ for expression in E. coli. T7 Promotor: The T7 promoter used to regulate gene expression of recombinant proteins. The T7 promoter is a DNA sequence 18 base pairs in length to the transcription start site at +1 (5' - TAATACGACTCACTATAG - 3') that is recognized by T7 RNA polymerase. T7 Terminator: The T7 terminator marks the end of a gene and leads to the termination of transcription. Strep-tag: A Strep-tag is a peptide that consist of 8 amino acids which originates from the bacterium Streptomyces avidinii. The Strep-tag has a strong affinity for Strep-Tactin, which is used for purification of the fusion proteins or in our case peptides via affinity chromatography. LCI: LCI is a 47 amino acid long sequence that allows binding of the fusion peptide to polystyrene, thus the peptide can be immobilized on microtiter plates. This is particularly important for determining the affinity with the AS III assay. Linker 17x linker fused to peptide to ensure flexibility and freedom of binding sequence without interference of connected fuselage. GOI: GOI, here "most common standard tag" ("MC") to be evaluated for use in lanthanoid recovery process form e wast bioleaching solution. MC-peptide with the following amino acid sequence "YIDTNNDGWYEGDELLA" was found in scientific literature [1]. The DNA sequence given here was derived from the before given peptide sequence and codon optimized for higher expression rates in E. coli.
This construct was designed for expression of peptides – the GOI – to be assessed for their applicability in a process of metal recovery from hetoregenous acidic bioleaching solutions.
Source
Sequences combined, GOI "most common standard tag" amino acid sequence obtained from [1], coding sequence derived from aa sequence and codon optimized for expression in E. coli. Vector containing BBa_K3013000 and BBa_I757014 obtained from supervisor at Schwaneberg Lab
References
[1]: L. E. Rosaleny, S. Cardona-Serra, L. Escalera-Moreno,J. Baldoví, V. Gołȩbiewska, K. Wlazło, P.Casino, H. Prima-García*, A. Gaita-Ariño*, and E. Coronado: Peptides as Versatile Platforms for Quantum Computing Phys. Chem. Lett. 2018, 9, 16, 4522–4526 DOI: 10.1021/acs.jpclett.8b01813.