How to assess PTM localization confidence?

[Munchic]

Hello, I am trying to compare confidence of phospho localization in a few DIA-NN settings. So far, I added the following additional options: --monitor-mod UniMod:1 --monitor-mod UniMod:21 --monitor-mod UniMod:4

A few questions:

1. What is the difference between "PTM.Site.Confidence", "Lib.PTM.Site.Confidence", "PTM.Localising" and which one should I use for my purpose?
2. When adding --monitor-mod, that score is automatically incorporated in the above metrics and there is no separate column for each modification's confidence?
3. In general, how would you suggest comparing DIA-NN outputs from two sets of parameters? E.g., searching 2 var mods vs. 3 var mods.

Thank you so much for your help and guidance!

[vdemichev]

Hi,

1. Lib. values are those recorded during the library creation stage. Whether it's OK to use them instead of run-specific ones probably depends on the experiment. PTM.Localising is not supposed to be used.
2. DIA-NN reports localisation confidence for all sites on the peptide combined.
3. Number of modified peptides at PTM.Q.Value < 0.01.

Best,
Vadim

[Munchic]

Thank you, this is helpful!

[sooheon]

Hi vadim, the docs now say that ptm-qvalue is "legacy", does this mean for phosphosite filtering i should rely only on lib.q.value and ptm.site.confidence, and not ptm.q.value?

[vdemichev]

Hi Sooheon,

Please just use Peptidoform.Q.Value instead of PTM.Q.Value.

Best,
Vadim