Exporting protein name from proteome FASTA file. jpbelair  2024-01-02 12:06
 

I have a list of 328 gluten / gliadin peptides found in the wheat proteome of 1,750 wheat proteins. I have a candidate list of 28 peptides with very good signal and fragmentation. The sample matrix is Human urine and I am capturing the gluten / gliadin peptides by immunoprecipitation using a series of mAbs A1, G12, and two custom-made mAbs. I have imported the FASTA file of the wheat proteins. I am putting together a list of candidate peptides that serve as a biomarkers for Celiac Disease. The peptides are found in multiple wheat species. I can see the peptides associated with multiple proteins when I go to Edit --> Unique Peptides... I've tried playing around with the report template without success. Does anyone know how to export the protein name(s) (and possibly more data) from the FASTA file? Copying the data from the window does not capture the column header information showing the protein name.

sp|A7XUQ5|AVLB5_WHEAT Avenin-like b5 OS=Triticum aestivum OX=4565 PE=2 SV=1
MKVFILALLALTATTAIAQLETTCSQGFRQYQQQQQPGQRQLLEQMRPCVAFLQQQCRPL
RMPFLQTQVEQLSSCQIVQYQCCQQLAQIPEQIRCHAIHNVVEAIMQQQSQQQRQERQQQ
AQHKSMRMLLETLYLMCNIYVPIQCQQQQQLGQQQQQQLQEQLTPCATFLQHQCSPVTVP
FPQIPVDQPTSCQNVQHQCCRQLSQIPEQFRCQAIHNVAEAIRQQQPQQQWQGMYQPQQP
AQLESIRMSLQALRSMCSIYIPVQCPAPTAYNIPMVATYTGGAC

All I need is the protein name "A7XUQ5|AVLB5_WHEAT Avenin-like b5".

The data was acquired on a Q Exactive Plus with DDA acquisition.

Skyline version 22.2.0.351 (28f9b9301), Windows 10 Enterprise.

Thank you, Jeff
 
 
Nick Shulman responded:  2024-01-03 07:44
I do not understand your question, but if you give us more information it might make more sense.

Could you send us your Skyline document? In Skyline you can use the menu item:
File > Share
to create a .zip file containing your Skyline document and supporting files including extracted chromatograms and background proteome etc.

Files which are less than 50MB can be attached to this support request. You can upload larger files here:
https://skyline.ms/files.url

There is a relatively new feature which offers a completely different way of finding duplicate peptides. You can use the menu item:
Refine > Associate Proteins
to move the rearrange the peptides in the Targets tree so that they belong to the proteins that they can be found in.

A new option "Create protein groups" was added a few versions ago (it seems to be present in Skyline 22.2). If you choose to create protein groups, then non-unique peptides will end up belonging to a protein whose name has a slash "'/" character in it, which you could filter for in the Document Grid.

I believe we have made improvements and fixed some bugs in this Associate Proteins dialog in recent versions, so you might find this works better in Skyline 23.1 or Skyline-daily.
-- Nick
 
Nick Shulman responded:  2024-01-08 10:17
Thank you for showing me your Skyline document.
I see that the "Associate Proteins" dialog does not work for you at all because none of the peptides that you are looking for are tryptic peptides.
Currently, the Associate Proteins dialog will only assign a peptide to a particular protein if the peptide could have been generated from that protein sequence using the enzyme that you have selected at "Settings > Peptide Settings > Digestion".

I will ask around and see whether it would be possible for us to change this behavior soon.
-- Nick
 
Nick Shulman responded:  2024-01-08 12:02
Jeff,

I have made a special build of Skyline-daily for you where the "Associate Proteins" dialog does not care whether the currently selected enzyme could actually generate the peptide in question.
You can install this special Skyline from here:
https://proteome.gs.washington.edu/~nicksh/SpecialSkylines/AssociateProteinsIgnoresEnzyme/

Note that this particular installer has not been signed, so Windows will probably try to stop you from running it, but you should be able to push the "More Info" button and then the "Run Anyway" button.
This special Skyline will appear on the Windows Start Menu as "Skyline Ignore Enzyme".
I believe it is possible to have Skyline, Skyline-daily and this special "Skyline Ignore Enzyme" installed on your computer at the same time.

Since this special Skyline is based on Skyline-daily, the document that it saves will be incompatible with Skyline 23.1, but you can use the "File > Share" menu item to produce a .sky.zip file which could be opened by Skyline 23.1.

When we eventually get this functionality into a real Skyline-daily, the behavior will probably be a little more nuanced, where the enzyme matters if there is at least one protein that could have produced that peptide, but, if the peptide cannot be found anywhere, then it will accept non-specific cleavage.

Hopefully this special Skyline will unblock you and allow you to move ahead with your research while we figure out exactly how this feature should work.
-- Nick