Welcome back to PyMole Biomonicles. In this video series we are going to see how to do molecular dynamics of protein, Eigen complex and windows operating system. So, in this part we are going to see how to solve it this system and how to add the ions and how to do the energy minimization also. You all know this is our working directory and we have already opened the command prompt ok. Now, we are going to change the directory. So, now we are in the working directory. First we are going to give this command. GMXedit.com-f- complex.grp-vnewbox.grr-bt.doddecavatron-d1.0. You all know the explanation for this command. If you do not know see my previous videos. So, give enter. Now, we can see the new box is created. So, you can see this is the newbox.grpil. Next to give this command this command is for salvation. So, give enter. You can see the new file is created for the salvation. The new file name is the solve.grv. You can see this is the solve.grv. That is all we have solvated the system. Next we are going to clear the screen. Next we need to download some mdp files for that. Go to the MD2 Torres website and you can see in this step 4 adding ions you have to download the mdp files. So, click on this one this link. I am going to copy this and save it as a new file. ions.mdp. I am going to open with an outpad plus plus and I am going to paste the file contents and hit control s and save the file. Similarly, go to this link step 5 energy minimization and you have to download this mdp file. Here is the link. So, this is the mdp file. So, copy this also and create a new file. And save this file also. You can see in this mdp file the total number of steps is 50,000 for the energy minimization and stop minimization when the maximum force is less than 10 kilojoule per mole. So, total is 1000 and you can change any of these parameters in the energy minimization mdp file and you can also change in some of the parameters in the ions.mdp also. But this is just a template mdp file. You can use your own mdp files. So, that is all. We have downloaded the two mdp files. So, one is the energy minimization mdp file. Another one is the ions.mdp file. Okay. Next, give this command. GMX grow mpp dash f ions.mdp dash c solve.grwo dash p topal.top dash o ions.dp r. So, this is the output binary file ions.tp r. So, give enter and then now ions.tp r file will be created now. As you can see there is an error invalid delimiter for file name in the hash include statement. Soil is topal.top in the line 24. So, the last line read is hash include 2ep.prm. So, this line we have included in the topal.top. So, we need to resolve this error. So, go to the topal.top file. Open this topal.top file and this is the line number 24. So, we need to enclose this file name inside the double quotation mark. Okay. Let us try this. First we shall try this and we see whether the error is resolving or not. Okay. And I am going to give the same command. GMX grow mpp dash f ions.mdp and the same as before. The output file is ions.tp r and give enter. Let us see what is going to happen. So, you can see the same address recurring invalid delimiter for the file name in the hash include statement file topal.top in the line 24.479. So, that is 2ep.itp. Okay. So, we need to enclose this also. So, we need to enclose this 2ep.itp in the double quotation also. Okay. So, go to this 24.000 line. Okay. Okay. You can see we have missed the double quotation here also. So, add the double quotation before and after the file name. Okay. So, I think that is all. We have included these two files. So, we have added double quotation for these two files. Okay. So, save this file and let us try to run the command again. The same command the output file is ions.tp r. Okay. I think the ions.tp r file is generated now. So, this is the ions.tp r. So, we have to enclose the file names inside the double quotations. Okay. So, after adding ions, we have to perform the energy minimization. So, I am not going to change any of the steps in the energy minimization of dp. So, this is just a demo. So, I am going to run as it is. I am going to clear this screen. So, after the generation of ions.tp r, give this command to generate the ions. Okay. And this and this command will generate solve ions.grow file and it will update the topology file. Okay. And it will add the appropriate ions whether it is sodium or chloride. Okay. To make it to make the system neutral. So, select a continuous group of solvent molecules. So, I am going to select group number 15. So, that is the solvent molecules. Okay. So, you can see there is six chloride ions are added. We have already seen in some previous step, the total charge in the system is six. So, so there are six chloride ions are added. Next, give this command gmxgrowmpp-f-em.mdp-c-solv ions.grow. So, this is the output of the previous command-p-topal.top. So, we are going to update the topology file and generate the output file of em.tp r. So, give enter. So, now you can see the em.tp r file will be created here. This is the em.tp r file that we have generated now. Okay. Next, give this command gmxmd run-v different em. Okay. So, this command will run this binary file em.tp r and generate some output files. Okay. So, give enter. Now, the energy minimization is happening. Now, you can see the energy minimization is completed in the 267 steps and the steepest descent converged to fmax in less than 1000 in 260 sound steps. Okay. So, that is all. We have performed the energy minimization. So, in this video, we have seen how to solve the system, how to add the ions and how to do energy minimization. So, these are all very basic steps. See you in the next one. So, thank you for watching.