Team:Saint Joseph/Composite Parts
Composite Parts
Composite Parts
We have three composite parts:
1. pET26b + wild type laccase
pET 26b (BBa_K2962001), is an expression vector used for expression of Laccase gene in E. coli SHuffle organism.
The wild type laccase is our enzyme as we mentioned it in basic parts. These two together is a composite part. We used this to make the expression.
2. pET26b + pelB.laccase
-
What is Pelb leader sequence?
The pelb leader sequence is a sequence of amino acids which, when attached to a protein, directs the protein to the bacterial periplasm, where the sequence is removed by a signal peptidase. The leader sequence consists of the 22 N-terminal amino acid residues.
-
How It works?
This leader sequence can be attached to any other protein (on the DNA level) resulting in a transfer of such a fused protein to the periplasmic space of gram negative bacteria, such as Escherichia Coli, often used in genetic engineering.
When we use pelB and laccase together, it is much easier to get the laccase from the periplasmic area. And after we used our expression vector for the expression.
3. pET26b + dsb.laccase
The disulfide bond stabilizes the folded form of a protein in several ways:
- It holds two portions of the protein together, biasing the protein towards the folded topology. That is, the disulfide bond destabilizes the unfolded form of the protein by lowering its entropy.
- The disulfide bond may form the nucleus of a hydrophobic core the folded protein, i.e., local hydrophobic residues may condense around the disulfide bond and onto each other through hydrophobic interactions.
- Related to 1 and 2, the disulfide bond link two segments of the protein chain, the disulfide bond increases the effective local concentration of protein residues and lowers the effective local concentration of water molecules. Since water molecules attack amide-amide hydrogen bonds and break up secondary structure, a disulfide bond stabilizes secondary structure in its vicinity. For example, researchers have identified several pairs of peptides that are unstructured in isolation, but adopt stable secondary and tertiary structure upon forming a disulfide bond between them.
These are the reasons why we used dsb+laccase. It is a method that we made to see if laccase works better with dsb or doesn’t. And then again we used our expression vector for the expression like we always do.
Saint Joseph Composite Parts
| Name | Type | Description | Designer | Length |
|---|---|---|---|---|
| BBa_K2962006 | Composite | pET26b with laccase enzyme from Trametes Versicolor. | Sera Küçük | 6943 |
| BBa_K2962007 | Composite | pET26b with pelB laccase (signal peptide laccase) | Sera Küçük | 5431 |
References
- [1] Mohamed Achraf Blout. Développement de biocathodes pour biopiles enzymatiques utilisant la laccase. Chimie théorique et/ou physique. Université Pierre et Marie Curie - Paris VI, 2017. Français. ffNNT : 2017PA066249ff. fftel-01695524f