Skyline not recognizing Heavy Acetyl modification of lysine residues

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Skyline not recognizing Heavy Acetyl modification of lysine residues dbade001  2019-09-20
 

Hey,
I am trying to upload my maxquant files to skyline. It has worked for my other files but it seems to have an issue with the heavy acetyl modification on the lysines. I am uploading a pic of this. Have you seen this issue before?

Thanks for your assistance and effort :)

David

 
 
matt.chambers42 responded:  2019-09-23

Hi David,

Can you attach your mqpar.xml and modifications.local.xml and modifications.xml files?

 
dbade001 responded:  2019-09-23

Hey Matt,
The version of skyline I was using doesnt seem to create the local file. I added the other two. I was working with two searches with unique mqpar.xml files, but both seem to have the same issue when I tried to isolate them.

Thanks
David

 
dbade001 responded:  2019-09-23

Hey Matt,
The version of skyline I was using doesnt seem to create the local file. I added the other two. I was working with two searches with unique mqpar.xml files, but both seem to have the same issue when I tried to isolate them.

Thanks
David

 
matt.chambers42 responded:  2019-09-23

It's MaxQuant that creates the modifications.local.xml file: Skyline will use it if it's present. I can't see why Skyline wouldn't be able to handle this as long as the modifications.xml file you attached was accessible to Skyline (i.e. in the same directory as msms.txt or parent or grandparent directory). If so, then can you also attach your msms.txt so I can try to reproduce myself?

In the meantime, can you try to upgrade to the latest MaxQuant? It uses a different way of specifying modifications that's less ambiguous than the old way. For example if you were searching with both Acetyl light and heavy mods at the same time, the old format couldn't handle it (the (ac) would be ambiguous) but the new format can.

 
dbade001 responded:  2019-09-23

Apologies, I meant the version of maxquant :) Ill add in the msms file.

Is there any risk of the modifications list in maxquant being deleted? I think that is the main concern behind why we havent updated yet.

Thanks for your help Matt!

Also, just to check because I was making the library from two searchs, ill reupload the mqpar file to make sure its the correct one.'

Also, I truncated this msms file to remove those peptides that Im not interested in.

 
dbade001 responded:  2019-09-23

Also, I was searching for both N-terminal 'light' acetylation and heavy acetyl modified lysine. Is there a chance it had issue with using a heavy modification?

 
matt.chambers42 responded:  2019-09-23

OK yes this second mqpar.xml would suffer from the ambiguity problem I mentioned earlier. You'll need to use the newer MaxQuant for this multi-acetyl search to work with Skyline. I recommend moving your custom mods to a modifications.local.xml file and let MaxQuant overwrite the modifications.xml with its new version. That way you'll keep both your local mods and get any new mods from central curation.

 
dbade001 responded:  2019-10-01

I updated maxquant and attempted in process my files having the heavy acetyl modification of lysine as a label.
The program says the label is not in the modifications file, but it is. Im not certain how to proceed on fixing that, have you seen this before?

Im attaching the msms file, mqpar, modifications.xml. The msms file was compressed with 7zip so I could send it.
I also added a screenshot of the error message.

 
matt.chambers42 responded:  2019-10-01

The msms and mqpar match but I don't think they're the right files. They have these mods:
<variableModifications>
<string>N-term Dimethyl</string>
<string>N-term Methyl</string>
<string>N-term Trimethyl</string>
</variableModifications>

 
dbade001 responded:  2019-10-01

I placed heavy acetylation as a label for this run, in the hopes that the issue involved it being a heavy isotope modification.
I also eliminated N-terminal acetylation as an option, to further remove possible problems.

 
matt.chambers42 responded:  2019-10-01

Yes, I'm able to reproduce that. Strange. I'll work on debugging that but in the meantime you can redo your search with the new MaxQuant and using the heavy label as a standard mod. That definitely SHOULD work. (I have very little experience with labels in BiblioSpec, but it looks like I'm going to get some now!)

 
matt.chambers42 responded:  2019-10-09

OK I think the issue is that you used "2H" in the composition instead of "Hx". Although MaxQuant does seem to work with 2H, its own labels use Cx, Hx, Nx for heavy versions of those elements. But since MaxQuant works with 2H, BiblioSpec should support it too.

 
dbade001 responded:  2019-10-09

Thanks for your effort. I'm giving it a try now and will let you know :)