Hi,
I am looking for a better software to analyse post translational modifications on the proteins after Mass Spectrometry.
I am purifying AMPylated (Adenylylated) peptides through a two step immunoprecipitation method using AMP(Adenosine monophosphate) specific antibodies. After tryptic digest, I have performed H-CID based MS/MS analysis on these peptides. My techniques is working fine, and I am getting peptides that should be AMPylated or phosphorylated, but I am not observing the modification in the software.
The peptides observed as modified have very low Mascot score and therefore, could not be considered significant. I am using Proteome discoverer 2.4 for analysis. I've tried Mascot too. I added phosphoadenosine (AMP) modification as a variable modification in these softwares. I even tried adding Phosphorylation as a variable modification and they did not show this modification on significant peptides that should be phosphorylated. Can someone guide me alternative methods/softwares to analyse these PTMs?
Please look at the following below links which may help you in your analysis:
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5548100/pdf/nihms888670.pdf
https://www.ncbi.nlm.nih.gov/books/NBK56012/
https://www.future-science.com/doi/pdf/10.2144/000112201
Thanks
Please check Unimod:
http://www.unimod.org/modifications_view.php?editid1=405
Narjis Fatima , Targeted proteomics methods (SRM/MRM/PRM) may be better if you have already planned targeted proteins. Otherwise, if you want to analyze by global proteomics method, SWATH or other DIA method may be suitable, I think.
https://www.youtube.com/watch?v=QryQe9TO97s
Alexandre Podtelejnikov Thanks for the link. I am using the same modification in MasCot as a variable mod, but I am unable to get any modification. May be the AMP residue is cleaved during Collision induced dissociation .
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