Highlights of DC-PPMA
(i) A major facet of DC-PPMA is that the user can‘view’ the database (in the form of an excel file) that they
will be using further for MS/MS based search or data analysis, viz.,
prior to database search, the user can know, what are the m/zvalues of precursor ions (of proteolytic/truncated peptide sequences)
and what are the m/z values of fragment ions that will be
involved in the search engine process. To the best of our knowledge,
this particular facet is not available in any proprietary or online tool
that are utilized for proteomic or peptidomic investigations. By
viewing/knowing the database containing m/z values of precursor
ions and fragment ions, it is possible to know better about ‘true
negatives and false positives’ and the user obtains a better
understanding about the process of matching between the experimental
MS/MS data and the theoretically calculated values present in the
database. Therefore, if DC-PPMA is used to construct appropriate ‘decoy
database’ by choosing suitable protein sequences, then he/she can obtain
better comprehension about false discovery rate (FDR). Thus, viewing and
analysis of decoy and target databases can be very helpful in more
reliable annotation of MS/MS spectra towards proper protein
identification.
(ii) Another notable feature of DC-PPMA is that the Module 1
calculates the m/z values for ‘singly as well as multiply charged
(protonated)’ precursor ions (viz., proteolytic peptides) and also
calculates the m/z values for ‘singly and multiply charged
fragment ions: a-, b-, c-, x-, y- and z- ions’. Consequently, in the
‘custom’ MS/MS database (created from Module 1), the m/z values
of each fragment ion (e.g., b, y, c, z ions) and their respective
‘charge state’ are generated and shown in the excel sheets, which can be
anticipated from MALDI and ESI MS/MS experiments. Therefore, DC-PPMA can
be useful for ‘manual spectral annotations and interpretations’ for
proteomic researchers and/or protein/peptide chemists, who use both ESI
and MALDI MS/MS.
(iii) Additionally, DC-PPMA can be of immense utility for
‘targeted analyses’, particularly for multiple reaction monitoring
(MRM ) based experiments, wherein it is essential to‘design suitable channels’ that should encompass the m/zvalues of precursor and pertinent fragment ions. In this context, the
output from Module 1 can be used for ‘designing MRM method’ ,
because the custom database built by Module 1 would consist ofm/z values of singly as well as multiply protonated precursor and
relevant fragment ions. Thus, DC-PPMA can be helpful for quantitative
studies also.
(iv) Furthermore, peptide sequences, either single or multiple
sequences can be uploaded in DC-PPMA. This particular feature can be
useful for peptidomic investigations and also for de novosequencing exercises or assignments. Consequently, DC-PPMA can indeed
prove to be worth for de novo sequencing of polypeptides and
proteins as well. Additionally, it can be used for discovery based
proteomic/peptidomic analysis also, if/when the transcriptome and/or the
genome of their sample is also known.
(v) In addition, the custom databases generated by DC-PPMA can
also be utilized further for planning about probability-based scoring
algorithms or scoring schemes, so as to identify more peptides and
proteins in a reliable fashion.