Team:Berlin/Modeling
7. Modeling
Modeling the S. Salmonella Typhimurium
After the design of a template Flagellin (fliC MCS) lacking the D3 domain and bearing a multiple coning site instead (by Johann) different D3 domain variants were designed for the purpose of our project. The aim of the modular D3 part design is to incorporate noncanonical amino acids either via SPI method or SCS method [1] in the variable Domain of Flagellin.
Furthermore, the addressable sites displaying the functional side chains of the noncanonical amino acids had to be identified and determined, too. For that our Jan (Bioinformatician) developed a pymol script giving all possible combinations of sites forming an equilateral triangle. A further filter parameter was given by the choice of exposed uncharged amino acids outside structural regions such as alpha helices and beta sheets.
An equilateral triangle (>25 Â) was chosen for the following reasons:
1) The D3 domain provides a very small surface in the shape of a hemisphere
2) Maximum distances between the functional side chains are of interest to avoid steric clash after functionalization
3) a maximum of three positions provides the larges possible functionalization.
In our investigations this scenario is provided by an equilateral triangle. However, other D3 domains bearing the information for incorporating two ncAA and one ncAA were also designed for means of comparison after functionalization with our enzymes of interest being the plastic degrading enzymes such as Cutinase.
Using the software pymol suitable residues were calculated to minimize steric clashes of the enzymes that will be attached to these residues via click reaction.
[1] Hoesl MG, Budisa N. Recent advances in genetic code engineering in Escherichia
coli. Curr Opin Biotechnol. 2012 Oct;23(5):751-7.
doi: 10.1016/j.copbio.2011.12.027. Epub 2012 Jan 9. Review. PubMed PMID: 22237016.
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