• For the respective proteins, We scavenge the RCSB Database for the Lactoferrin and Amyloid-Beta Protein Structures, Picking 1LFG(Human Lactoferrin-Diferric) and 1IYT(Amyloid Beta -42, Monomeric) and 5KK3(Amyloid Beta-42, Fibril Structure)
  • Aptamer for Lactoferrin obtained from iGEM repository, while a list of DNA-Aptamers for Amyloid beta obtained from Literature review.

  • We obtain the 2-D structure of the DNA-Aptamers using the MFold Server.
  • Inputting the files into the Xiao Lab web server to obtain the 3-D structure of Protein in .pdb format.
  • Using UCSF Chimera, we process the Protein and Aptamers, for removal of non-standard residues in the structures including ions, glycosides, and water molecules to prepare for docking.
  • Saving the processed file in the .pdb format

  • We perform Docking using HADDOCK 2.4 web server, selecting the option for Protein-Nucleic Acid docking, uploading the required input files
  • We select all residues for docking, and select the Centre of Mass restraint for docking, while keeping the default parameters.
  • The results for each Protein-DNA Aptamer docking is obtained in the form of Clusters associated with a HADDOCK Energy. Each cluster has 2-4 structures.
  • Choosing the Cluster with lowest HADDOCK energy, and downloading all the structures In .pdb format associated with the cluster for MD runs.

  • Using the Solution Builder tab on CHARMM-GUI web server, firstly inputting the docked Protein-Aptamer structure obtained from HADDOCK, and manipulating the residues in case of any problems.
  • Then We in-silico generate a Rectangular WaterBox with edge distance of 10 Å, and add 0.15 M KCl in the solution, using the ion-placing method called Monte-Carlo, The Water Module used is TIP3
  • We generate the Crystal type Box(Cubic), with the parameters, now automatically generating grid information for PME FFT.
  • Now we generate the Force Field files and input files for MD-Simulation, selecting CHARMM36m force field and GROMACS MD-software for input files generation
  • Equilibration methodology used is NVT Ensemble and Dynamics methodology used is NPT Ensemble with Temperature: 303.15 K
  • Download the zip file for, energy minimization, NVT and NPT input files along with a gromacs file of Protein-Aptamer in Solvation box along with topology file showing the coordinates of the atoms.

  • Firstly we determine the simulation time by changing the nsteps parameter in the above mentioned files, For Lactoferrin( 50 ns) and for Amyloid Beta-42(100 ns) have been determined.
  • Energy Minimization - The First step of the simulation with n_steps

Formula for nsteps: nsteps * timestep (ps/step) = 1000 * x ps = x ns

  • NVT Equilibration - Nsteps according to the Simulation Time
  • NPT Dynamics
  • MD Run Final

  • Recentring and Re-wrapping the coordinates -
  • RMSD Calculation -
  • RMSF Calculation -
  • H-Bonds Calculation -