Engineering dynamic gates in binding pocket of penicillin G acylase to selectively degrade bacterial signaling molecules

<p>(01-mutants_design.tar.gz) Mutants design:</p> <ol> <li>Input structures of ecPGA from the PDB database (PDB IDs: 1GK9, 1GM7, and 1GM9), processed to resemble wild-type state, repaired by RepairPDB module of FoldX&nbsp;4</li> <li>Double-point mutants preparation, analysis and filtering: <ol> <li>text files including configuration for FoldX 4</li> <li>inputs and outputs of CAVER 3.02 calculations on FoldX 4 PDB files of ecPGA double-point mutants</li> <li>input configuration file and TransportTools library 0.9.4 calculations outputs generated based on the inputs produced in step 01 and 02 above</li> <li>CSV files containing complete information about FoldX 4 stability prediction and geometrical properties from CAVER 3.02 and TransportTools library version 0.9.4 for ecPGA double-point mutants</li> </ol> </li> <li>Triple-point mutant preparation, analysis and filtering: <ol> <li>text files including configuration for FoldX 4</li> <li>inputs and outputs of CAVER 3.02 calculations on FoldX 4 PDB files of ecPGA triple-point mutants</li> <li>input configuration file and TransportTools library 0.9.4 calculations outputs generated based on the inputs produced in step 01 and 02 above</li> <li>CSV files containing complete information about FoldX 4 stability prediction and geometrical properties from CAVER 3.02 and TransportTools library version 0.9.4 for ecPGA triple-point mutants</li> </ol> </li> </ol> <p>(02-docking.tar.gz) Preparation of protein-ligand complexes using molecular docking for wild-type ecPGA and 6 best designed triple-point mutants with 6 various bacterial signaling molecules:</p> <ol> <li>PDB files of ligand, PDBQT files of the receptor and PDB files of the complexes selected from docking experiment:</li> </ol> <p>Full names of presented protein variants:<br>ecPGA_wt, wild-type Escherichia coli penicillin G acylase<br>LAF, Phe138&alpha;Leu &amp; Met142&alpha;Ala &amp; Ile177&beta;Phe ecPGA variant internally referred as 1GK9_Repair_22<br>LSF, Phe138&alpha;Leu &amp; Met142&alpha;Ser &amp; Ile177&beta;Phe ecPGA variant internally referred as 1GK9_Repair_98<br>MAF, Phe138&alpha;Met &amp; Met142&alpha;Ala &amp; Ile177&beta;Phe ecPGA variant internally referred as 1GK9_Repair_23<br>MSF, Phe138&alpha;Met &amp; Met142&alpha;Ser &amp; Ile177&beta;Phe ecPGA variant internally referred as 1GK9_Repair_99<br>VAF, Phe138&alpha;Val &amp; Met142&alpha;Ala &amp; Ile177&beta;Phe ecPGA variant internally referred as 1GK9_Repair_30<br>YAF, Phe138&alpha;Tyr &amp; Met142&alpha;Ala &amp; Ile177&beta;Phe ecPGA variant internally referred as 1GK9_Repair_33<br>Full names of presented AHLs:<br>C06, N-hexanoyl-L-homoserine lactone;<br>C06-3O, N-3-oxo-hexanoyl-L-homoserine lactone;<br>C08, N-octanoyl-L-homoserine lactone;<br>C08-3O, N-3-oxo-octanoyl-L-homoserine lactone;<br>C10, N-decanoyl-L-homoserine lactone;<br>C12-3O, N-3-oxo-dodecanoyl-L-homoserine lactone</p> <p>(03-protein_ligand_MDs.tar.gz) Ligand-enzyme complexes molecular dynamics for wild-type ecPGA and 6 best designed triple-point mutants with 6 various bacterial signaling molecules:</p> <ol> <li>Force field parameters in Amber format</li> <li>Input coordinates *.inpcrd, parameters *.parm7 and *.pdb files for each complex ready for simulation in Amber</li> <li>Amber input files *.in for minimization, equilibration and production runs</li> <li>Restart files for each stage of the minimization, equilibration and production runs in Amber *.rst format</li> <li>Simulation output files for each stage of the minimization, equilibration and production runs in Amber *.mdout format</li> <li>Output files generated during post-processing of production runs trajectories in a form of text files generated by cpptraj</li> </ol> <p>(04-free_enzymes_MDs.tar.gz) Free enzymes molecular dynamics of 3 best triple-point ecPGA (VAF, YAF and MSF) mutants prioritized based on protein-ligand molecular dynamics simulations and experimental assays:</p> <ol> <li>Force field parameters and input coordinates *.inpcrd, parameters *.parm7 and *.pdb files for each complex ready for simulation in Amber format</li> <li>Amber input files *.in for minimization, equilibration and production runs</li> <li>Restart files for each stage of the minimization, equilibration and production runs in Amber *.rst format</li> <li>Simulation output files for each stage of the minimization, equilibration and production runs in Amber *.mdout format</li> <li>Post-processing analysis of generated trajectories: <ol> <li>Text files with distances, CSV files containing result of PCA and clustering, PNG files with clustered PCA results</li> <li>Inputs and outputs of MDpocket analysis and visualization of the pocket frequency grid as an isomesh</li> <li>CAVER input configuration files in text format, CAVER output data including parsed CSV and text files for visualization of entrance opening time evolution and cavity profiles inspection</li> <li>cpptraj generated text files including RMSD, distances and chi1 angles measurements</li> </ol> </li> </ol> <p>All plots were generated using matplotlib or seaborn Python libraries. Figures containing structural representations were generated using PyMOL 2.0.1.</p> <p>(05-ecPGA_VAF_YAF_MSF_penG_MDs.tar.gz) PenG-enzyme complexes molecular dynamics for wild-type ecPGA and 3 best designed triple-point mutants (VAF, YAF, MSF):</p> <ol> <li>PenG force field parameters in Amber (GAFF) format</li> <li>Input coordinates *.inpcrd, parameters *.parm7 and *.pdb files for each complex ready for simulation in Amber</li> <li>Amber input files *.in for minimization, equilibration and production runs</li> <li>Restart files for each stage of the minimization, equilibration and production runs in Amber *.rst format</li> <li>Simulation output files for each stage of the minimization, equilibration and production runs in Amber *.mdout format and analysis output files generated during post-processing of production runs trajectories in a form of text files</li> <li>Reactive Stabilization Score [RSS] statistics summarized in CSV files</li> </ol>