The idea of using skyline for crosslinked peptides using DIA in UDMSE mode, was because I read an application note from Waters Corporation called: "acquisition mode characterization for the qunatitative and qualitative analysis of cross-linked peptides by target and untarget LC- IM-MS. " and that it uses Skyline for the assignment of the ionic series; although I think they used ETD fragmentation. I know that to perform XL-MS, the workflow is done in DDA mode, but since I work in DIA mode (I feel that it is very robust to perform Label-Free) I thought that maybe the current versions of Skyline already support it and I was excited because they combined the way of working that I usually work in the lab with something that I want to exploit. My idea is to clone 2 proteins of my interest to interact and cross-link them with Disuccinimidyl suberate (DSS), enzymatically generate the cross-linked peptides to be analyzed. Using Ion mobility I can generate charge dependent clusters, and extract the mobility information of the ions of interest, in this case use peptides with charge state z = 3 + or higher; Then I can create a method in UDMSE mode to apply quasi-specific collision energy to crosslinked peptides and gain selectivity. What I would like skyline to do is to deconvolve the ions to be able to assign the ion series of the cross-linked peptides, in a other words, to know which peptides of the "X" and "Y" protein are cross-linked.
How long will it take to release the version you are telling me about?
Cheers! |
On that poster, it looks like they have told Skyline that these crosslinked peptides are "small molecules", where they have told Skyline the chemical formula of the intact molecule, and all of the fragments to look for. That certainly works today. It looks like the poster might be saying that they have some python scripts that they wrote to help with calculating the fragment ions.
We might release a Skyline-Daily with some crosslinking support in about a month. Skyline 20.2 will be the next official release of Skyline, which will probably happen late in the summer.
-- Nick |