How to find a suitable library for a certain protein (with enzyme digestion)

HENG

2020-04-10 06:49

Hello.

Recently I am analyzing a protein called leptin (Protein Name: P41159) with AspN digestion using Skyline. But even download some MS/MS spectral library form PeptideAltas and NIST. I cannot find a suitable library that match the transition of my peptides.

So whether there is a pathway that I can find certain library can match the transition of my peptides, when I input my target proteins and digestion enzymes in the searching field. Or other suggestion maybe you can give to me.

Thank you very much.

Heng

Nick Shulman responded:	2020-04-10 11:11
Skyline version 20.1 can use a web service called "Prosit" to predict the theoretical spectra of peptides. The way that you turn on this feature is go to: Tools > Options > Prosit and then choose something in the "Intensity Model" dropdown. After you have done that you can build a spectral library (.blib) file for yourself containing the predicted spectra of the peptides in your document. To do that, go to: Settings > Peptide Settings > Library > Build and for the "Data Source" option choose "Prosit". I am not sure that I understood your question. If your question was about something else, it might help if you could send us a screenshot of what you are looking at. -- Nick

Brendan MacLean responded:	2020-04-12 14:51
I feel I should note here that Prosit peptide spectrum prediction was likely trained with tryptic digested peptides and may not work well with arbitrary peptide sequences, or those created by AspN digestion. However, Skyline still has multiple options for you to create your own library. You could collect DDA data that includes your peptides; run a peptide search engine, and have Skyline build a library for the peptides that can be found with DDA using the Skyline library build mechanism described in the Targeted Method Editing tutorial (https://skyline.ms/tutorial_method_edit.url) You could collect targeted runs with PRM and export a library from Skyline for the precursors you feel convinced are detectable (either again through a peptide search or by injecting peptides using standard addition or stable isotope-labeled standards, or even high presence of the y-ion series coeluting), using File > Export > Library. Hope these are useful suggestions to you. I am not so sure Prosit would be useful for you in this case.