20221125 ProSight Proteome Discoverer Top Down Proteomics

ProSight PD is one of the most commonly used MS/MS identification algorithms for top-down proteomics. In this video, we walk through the steps required to identify proteoforms in a top-down experiment:
1) Acquiring a UniProtKB reference proteome in XML format
2) Importing the reference proteome and adding variable modifications
3) Setting up for a truncation search with 1% FDR control
4) Understanding the run-time information provided by ProSight PD
5) Interpreting the pdResult file tables in Proteome Discoverer.
Raw data from PXD019247 would be a good match for the E. coli lysate data I employed in the video, if you would like to reproduce this on your own computer.

[@samsam-kz4nf]

Hi, very nice video, i would like to ask about what your options toward maxquant and PD, is there large difference between them?

[@gabriellenna4895]

Hello David, it is an excellent video for me! I am new to the top-down field. The top-down approach is not well fragmented as the bottom-up and I would like to know why we choose the Uniprot database as the top-down search space? And is there any difference when we need to choose the search space for top-down and bottom-up?😃

[@DavidTabb]

Thanks, Gabrielle. The UniProtKB was chosen in support of top-down identification because it included considerably more information than just a sequence for each accession. By detailing different isoforms for each protein-coding gene, specifying amino acid substitutions, and enumerating well-documented post-translational modifications, UniProtKB annotates protein sequences more completely, making the recognition of an intact proteoform much more likely than considering the bare, wild-type sequence alone. With bottom-up, there's likely to be some wild-type short peptide matching the canonical sequence to enable ID, so all that added annotation is less critical.

[@rameshgunturu6120]

Dear sir i am working with orbitrap q exactive can we use run raw file prosite pd may i get your mail id need your help