Dear Skyline Team,
I would like to do DDA with MS1filtering in order to quantify a certain yeast peptide (TGAPNNGQYGADNGNPNGER) which is present in charge 2+ and 3+ with either dimethyl light or dimethyl heavy label in my dataset. I set the respective modifications in the Peptides settings tab and when I import my fasta file (which also contains some high abundant peptides to allow for matching between runs) I can see the expected m/z values present in my msms.txt (generated from MaxQuant Version 220.127.116.11). However when I next do the "import peptide search" all peptides vanish from the left side tab. When I go on I get the error message "The document does not contain any peptides". When I then try to add the Fasta file (again) I get the error message "The document must contain at least one precursor transition in order to proceed". After reading some of the earlier support threads I put the modifications.xml file from MaxQuant in the same folder (along with msms.txt and mqpar and Raw Files). I also tried the import replacing the Skyline names by the respective MaxQuant names, i.e. "DimethylLys0" instead of "Dimethyl (N-term)" in the peptide settings in Skyline. Still, Skyline somehow does not recognize the dimethyl modification from the msms.txt. When I try the import with default settings, Skyline suggests that there is Dimethl (K) my data and also an unkown K modification with the mass of 34.0 which corresponds to the heavy Dimethyl (K), but when I add them, I still get no transitions (but of course the respective N-terminal modifications are missing). When I try the import with just the Dimethyl (K) and Dimethyl (N-term) set as variable modifications (no heavy label), Skyline starts the import, however the peptide I am interested in is then only present as in the non-modified version. I tried all this with both the Skyline version 18.104.22.16819 and the Skyline version 22.214.171.124796. I would also like to add that the double dimethyl labelling settings I tried out now have worked before for another double dimethyl labelling dataset, which was also searched with the MaxQuant Version 126.96.36.199 but searched against a Trypanosoma database. I attach my msms.txt, mqpar and skys file in the hope that you can find out where the error is.
Dear Skyline Team,
I have found a workaround for my problem. I set both the light and the heavy dimethyl as structural, variable modifications. During the "Import peptide search" Skyline suggested that there is an unknown modification with the mass of 34.0 on each amino acid. I then added the heavy modifications on K and Nterm (giving them a different name) additionally as heavy modifications (mass shift of 6). The import started and I now get my peptide in the heavy and light version.