Quoting Youbin Tu ytu@mix.wvu.edu:

> Hi Alicia, Ben: > Thanks for your reply, which help me a lot. > But for my specific situation, the ligand I am trying to use is > phenacetin which did not exist in the template pdb file. Or to say, we > created it so as to make it docked in the active site by using some NMR > restraints. We expected to see some interactions between the ligand and > the active site which will make the active site show a differnt look > from what it is when no ligand is bound.

You have used NMR perturbation data to dock a ligand into the active site? You could simulate a ligand-induced conformational change in the receptor just by doing some MD on your complex. I recommend NAMD. However, AMBER will probably work better for you since your compound has an aromatic ring.

> Now I was really confused, if the ligand can not have correct > connectivity, how can we expect to see the interactions between the > ligand and the enzyme? I mean how the follwong refinement steps which > is involved with minimization and molecular dynamics correctly dealt > with the ligand and the interaction between the ligand and the enzyme?

MD packages rely on complete topology/parameter sets. You can use XPLO2D to predict a lot of this information. However, be aware that the values are completely derived from the pdb file and will need to be edited. Charges will also need to be added.

> Again, why the ligand does not show correctly, you know , we build > the phenacetin in Builder module in InsightII and output it as a RTF > file, then append it to the top_heav.lib file. The molecule looks very > reasonable in InsightII, but after calculation by MODELLER , the ligand > looked weird. So, any other program can view RTF file so as to have a > check the connectivity of the ligand is reasonably good or not? Or if > the parameters are incomplete, how to make it complete?

Although we have InsightII, I have never built an RTF file with it. I'm guessing it works similar to XPLO2D. Therefore you will most certainly have to edit angle and charge terms. However, you'd think that bonding would be correct.

> Another question is whether does the RTF file provide enough > connectivity information for the ligand? Or is it because of the > special restraints we applied in MODELLER which make the connectivity > changed? You know,we even tried to use the ligand without any > restraints, the result still looked weird?

To check if your RTF file provides correct connectivity, pass it through X-PLOR along with your pdb file to produce a psf. You can then view your psf file in VMD to make sure your connectivities are correct. Otherwise just go through the topology manualy.

> Basically, we just want to build a new ligand with some experimental > restraints in the active site of a enzyme. If the ligand looked werid, > we have to doubt whether the interaction between the ligand and the > enzyme is reliable or not? So, we have to know which step create this > problem?

If you already have the structure of the protein you are docking to then I wouldn't recommend using MODELLER to dock your ligand and observe ligand-induced confomrational changes. Do MD.

> 1. building the ligand (RFT file)? We did not give enough connectivity > information? > 2. running MODELLER? Since there is no restraints to confine the bond > or angles for the ligand or the restraints are conflicting with each > other which make the ligand broken? > 3. outputting file from MODELLER? Assumed that all the above is good, > the only problem is that MODDLER only give the cooridinates of the > atoms instead of the relationship ( connectivity) between atoms? > Hope I have well explained my problems? And I also attached my RTF > file for phenacetin. > Finally, thanks for your guys' kind help. Anyone's reply in this > topic will be highly appreciated. > Hope for the best. > > youbin >

Sorry I didn't go through your RTF file but its laborious work even with the smallest of compounds. Best create the psf file and then visualize it in VMD.