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Getting the structure of Alyteserin-1a

Alyteserin-1a is a modified antimicrobial peptide which was made by introducing two mutations (D4E & N23S) in Alyteserin-1c peptide. The structure for Alyteserin-1a is not available therefore we obtained the pdb structure for Alyteserin-1c (pdb id : 2L5R) and created the given mutations in pymol. We carefully analyzed the structural feature of Alyteserin-1a peptide to come up with another peptide which could interact with it. We seem to find an interesting pattern which was a hydrophobic groove on one side of the peptide. We targeted this region to neutralize the anit-microbial peptide.

Predicting the structure of Naly

In our project, a key step was to neutralize the antimicrobial peptide, Alyteserin-1a, so that we can create a stress free environment. For this, we designed a novel peptide of smaller length, which can interact favorably with Alyteserin-1a. We used following step to get the best possible peptide :

• We used pepstr, an online tool, which takes amino-acid sequence as input and gives us possible structures, in pdb form, of the protein.
• After getting several possible structures, we used another online tool, ZDOCK, which docks two molecules and give a score, which tells us about that how favorably peptides are interacting.
• We screened few peptides from above process and proceeded for MD simulations.

Molecular Dynamics Simulation

We performed molecular dynamics simulation experiments to confirm that Naly is able to interact with Alyteserin-1a antimicrobial peptide, hence it can neutralize it's activity to kill the pathogemic bacteria. Following are the steps that we followed in this experiment :

1. Convert both the pdb files to .gro files and get the topology files using pdb2gmx tool. Use GROMOS96 53a6 force field.
2. Change the positions of both the peptides in a way that they are far away and disoriented.
3. Build the .gro file for the complex (both proteins) by merging the above .gro files. Create the .top file for complex both peptides.
4. Define the box for the complex and solvate with water molecules.
5. Add NA+ and CL- atoms to neutralize the charge of whole system.
6. Prepare the .tpr file for energy minimization step.
7. Steepest descent minimization algorithm was used for a maximum of 50,000 steps.
8. Position restrain the water molecules in complex.top file.
9. NVT equilibrate the system.