Part:BBa_K5073030:Design
L7Ae-CD63
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal XhoI site found at 620
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
The following design points need to be considered when designing the sequence for L7Ae binding to the C/D box. Firstly, the L7Ae protein is highly specific to bind to the kink-turn RNA structure in the C/D box, so the sequence and secondary structure of the C/D box must be accurately designed to ensure efficient recognition and binding. At the same time, the sequence of the C/D box needs to be kept stable to enhance its expression efficiency and structural integrity within the cell. The physical space requirements of the L7Ae and C/D box also need to be considered during design to ensure that the anchoring process does not interfere with other molecules or functional elements. In addition, if the L7Ae or C/D box is designed to be part of a fusion protein (e.g., fusion with CD63), it must be ensured that the fusion does not affect the RNA-binding function of the L7Ae or the structure of the C/D box. Finally, codon optimization of the sequence was performed to ensure efficient expression and functional execution in different cell types.
Source
The L7Ae portion of the L7Ae-CD63 construct originates from the Archaeoglobus fulgidus ribosomal protein, which specifically binds to kink-turn RNA structures. The CD63 portion comes from the human CD63 gene, which encodes a tetraspanin protein commonly found on the membranes of exosomes. This fusion protein is created synthetically by combining the L7Ae and CD63 sequences to exploit their respective RNA-binding and exosome-targeting functionalities. Thus, the part is a synthetic construct combining elements from both archaeal and human genomic sequences.