Feature Request Sequence Dependent Precursor Charges

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Feature Request Sequence Dependent Precursor Charges roman sakson  2019-06-13
 

Hello everyone,

first of all, thanks again for the amazing piece of software that continues to be the highlight of my research days!

I work in a core facility, where we routinely develop MRM assays from crude synthetic standards without much prior knowledge, this means that for the first runs we have to "guess" the precursor charge state as well as suitable transitions based on common MS sense for relatively long target lists. In our experience, doubly charged precursor ions work best for tryptic peptides of average length, therefore this is the standard transition setting. However, if the sequence contains histidine, normally the more intensive precursor is triply charged. Is it possible to specify in the filter settings or elsewhere, that precursors should be doubly charged, unless there is a particular amino acid present in the sequence?

Thanks a lot in advance for your time!

Roman (Heidelberg, Germany)

 
 
Tobi responded:  2019-06-17

Hello all,

if possible I would like to add something which has also been mentioned before. It would be nice to have an automated charge state refinement. The idea would be (for standard peptides etc.) to build chromatograms for charge states 2+ to X+, manually check and correct peak picking, and with a few clicks to filter the target list to have only the most intense charge state left over. Removing the need of having to make deletions for every single peptide would be a big plus.

Best regards,
tobi

 
roman sakson responded:  2019-07-03

Dear Skyline team,

I just wanted to ask again since it has been a while since Tobi and I posted to this thread and I thought that it might have been overlooked.

Thanks,

Roman

 
Brendan MacLean responded:  2019-07-12

A summer intern, Padyoth Vemulapati, has started work on the feature for automatically refining large lists of peptides with more than on precursor charge to on the charge state with the most intense peak.

https://skyline.ms/issues/home/issues/details.view?issueId=664

Another intern, Tobi Rohde, is working on integrating Skyline with the Prosit deep-learning prediction tool, which currently predicts MS/MS spectra and retention times, and we hope Tobi will also get a chance this summer to make it predict best precursor charge state.

We don't currently have plans to implement a way for users to tell Skyline about heuristic precursor charge state prediction methods (like containing a Histidine). However, it has been possible for a long time to specify charge state in both Edit > Paste and Refine > Accept Peptides. That is, if you paste:

PEPTIDER+++

Into the Targets view, you will get the peptide with only the charge-3 precursor. While, if you have the peptide PEPTIDER in Skyline with say charges 1, 2, 3, and 4, and you enter only the above line into Refine > Accept Peptides, this will leave you with only the charge-3 precursor in the document, removing the other 3 charge states.

Nick Shulman also implemented a while ago the ability to delete targets from Document Grid, which would allow you to sort and filter a precursor report, select the ones you want to delete in bulk and click the Delete button (red X) in the Document Grid to get rid of them.

Hopefully, these features are or will be useful enough to get by on. Thanks for giving us your feedback!

--Brendan

 
roman sakson responded:  2019-07-12

Hello Brendan,

thank you for answering, for creating the max precursor issue and for the explanations! We normally prefer to import FASTA files rather then ready peptide lists. I could use the search function to look for peptides in my document that contain "H" (in the case of Histidines). The "Find Results" window that appears is very helpful to navigate the document! However, it would help me even more if I could then sort the hits in the "Find Results" window, as it is already possible for other grids in Skyline (for instance, according to type, so that all the Peptide-category hits would get grouped together to quickly go through them). Is it something that you might consider implementing in the future or do the results hits have to be in the order in that they appear in the target list of the document?

Thank you,

Roman

 
Brendan MacLean responded:  2019-07-12

Hi Roman,
I think you will have a much easier time achieving this with the Document Grid. It has a Contains filter. Just

  • View > Document Grid and from the Views (4.2) or Reports (Skyline-daily) menu, choose Precursors.
  • Click on the "Modified Sequence" column header and choose Filter.
  • Add Filter type: Contains and H.
  • Click OK.
  • Then do the same for "Precursor Charge" Equals 2.

Now you have a list of all peptides containing H and their charge 2 precursors.

  • Ctrl-A to select everything.
  • Click the delete button (red X) in the Document Grid toolbar

Skyline should ask if you want to delete N precursors.

  • Click OK

I just did this on a large document where N was over 10,000 (I believe including decoys. So, I would probably remove the decoys before doing this next time). Skyline gave me no progress and took a while to complete the operation (hopefully Nick can improve that), but it did work.

This is how I would advise you to go about bulk deleting targets from your document. Much easier than using Find and actually selecting everything you want to delete in the Targets tree.

Hope this helps.

--Brendan

 
roman sakson responded:  2019-07-12

Hi Brendan,

thank you very much, this is way better than what I had in mind! Also, I definitely need to have a closer look at the different filters that are already available, I barely make use of them at the moment.

Best regards,

Roman