Friday, December 12, 2014
Preprocessing TMT spectra for better IDs
This is the paper I mentioned earlier that introduced me to TurboRaw2MGF. This is one of those papers where you think, "this is already published, right?" Turns out it isn't.
Back up, Ben! Okay.... When we TMT or iTRAQ tag something and fragment it the reporter ions come off. Thats how we get quan. Unfortunately, Sequest looks at those fragments and doesn't know (by itself) that those aren't peptide fragments.
Turns out that if you remove these fragments from consideration, the search engine doesn't have to think about them and everything works better. In fact, there is a feature in Proteome Discoverer (don't ask me which one, its after 5 on a Friday here and I'm totally jetlagged....) that will take these out. This paper shows how much better your IDs can be if you deconvolute the MS/MS spectra and take out these reporter ion artifacts.
Worth checking out if you do these kinds of studies. Maybe it seems kind of obvious, and maybe you've already been doing this, but I doubt it would have published in MCP if you'd submitted it already! You can find it (currently open access) here.