Team:Virginia/Model
We modeled our system in silico to select a sterically feasible protecting group and to optimize a mutant leucyl-tRNA synthetase for complementarity of its catalytic site to protected leucine, and of its editing site to leucine. To select a protecting group, the team used protein-ligand docking software to compare binding affinities of several protected leucine/synthetase complexes. To perform mutagenesis on leucyl-tRNA synthetase, an integrated software script was written in the Linux shell, with inputs including a protein to mutate, a ligand, a list of residues of interest, and binding pocket location. The script runs mutagenesis, assesses mutant protein stability, then performs ligand docking. The program then ranks the outputs, acting as a streamlined mutagenesis optimization algorithm. We confirmed, using CSM software suites and iGEMDOCK, that AMP and AMS yield energetically comparable binding affinities. Lastly, we performed Michaelis-Menten modeling for the enzyme pepsin to gauge activity in nonspecific cleavage enzymes.
Leucine Synthetase SelectionSynthetase File: 4aq7
The Leucine tRNA Synthetase used in the following simulations was taken from the RCSB protein databank. The file used was 4aq7, a leucyl trna synthetase in its aminoacylation conformation.
AMS vs AMP
Protecting Group
Selection - Modeling Considerations
Several programs were used to asses the viability of the different protecting groups in our system. The main programs used for this purpose were Autodock Vina, iGEMDOCK, and CSM-Lig.
Residue Selection
Literature
Autodock Vina and Ligplot
Final Selections
MUT
Purpose
Components
Output
Source
MUT on GitHub Pepsin Modeling
Purpose
Results
Conclusions
