[Chimera-users] Help in validation of Mutant Structure

[Elaine Meng]

Hello Carlo,
Swapaa only performs a simple sidechain replacement — it does not change the backbone or predict any other changes in the structure that might occur due to the mutation.  Matchmaker RMSD uses only the backbone alpha-carbons (CA atoms), so the value of 0.0 after swapaa is expected.  Minimization will not really predict the change either.  It will only relax to the nearest local minimum conformation (not find the global mininum), typically not much different.

Your subject line “validation of mutant structure” is not what swapaa does.  Instead you have to use your own knowledge of protein structure to figure out if the mutation is disruptive.  In Chimera you can try to see if sidechain contacts, hbonds, etc. change. Instead of using the swapaa command, which automatically tries to pick the best sidechain rotamer, you can explore the mutation using the graphical Rotamers tool (in menu under Tools… Structure Editing) for better insight into effects on contacts and hbonds.  Choosing a rotamer will still only change the sidechain, however, not the backbone or other residues.
<http://www.rbvi.ucsf.edu/chimera/docs/ContributedSoftware/rotamers/framerot.html>

For example of using Rotamers, see the structure analysis and comparison tutorial
<http://www.rbvi.ucsf.edu/chimera/docs/UsersGuide/tutorials/squalene.html>

I don’t know what steps in Chimera you used to “perform multiple sequence alignment.” If you have sequences in an alignment and the structure is associated with one sequence, using swapaa to “mutate” the structure does not automatically change the sequence.  Maybe that is what you tried to do.

To get a sequence with the change from swapaa, you would first have to use swapaa on the structure, and then open the sequence from that modified structure. For example, to add it to your sequence alignment, you would use sequence alignment menu: Edit… Add Sequence, and then choose From Structure and then the modified structure chain in that dialog.  Or, to open the modified sequence by itself, main menu: Favorites… Sequence, choose the modified structure chain in that dialog.
<http://www.rbvi.ucsf.edu/chimera/docs/ContributedSoftware/multalignviewer/framemav.html>
<http://www.rbvi.ucsf.edu/chimera/docs/ContributedSoftware/multalignviewer/multalignviewer.html#add>

I hope this helps,
Elaine
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Elaine C. Meng, Ph.D.
UCSF Chimera(X) team
Department of Pharmaceutical Chemistry
University of California, San Francisco

> On May 19, 2019, at 2:50 AM, carlo cast <Karlo96 at hotmail.it> wrote:
> 
> Good Morning to everyone, it’s my first time here writing at you, I hope I’m doing right.
> The point is that I’m working with a wild type enzyme from which  I got mutant versions by using swap aa command. My goal is to look at the effects that mutations have on the structure by superimposing the mutant form with the wild type one. When I perform matchmaker command, RMSD is equal to 0 and the structures overlap perfectly; thus I thought to minimize structure before superimposing and in this way I see a little difference in RMSD but it could be due to  the different steps  of the minimize process (e.g. adding protons) than the actual effect of mutations. Moreover when I perform multiple sequence alignment, the amminoacids I swapped actually are not different from the Wild type ones. If I point the cursor on the swapped aa I see the difference in name and side chain but by doing multiple sequence alignment they do not appear.  
> Any suggestions?