Mass Spec data associated with Proteomics analysis of the polyomavirus DNA replication initiation complex reveals a novel functional phosphorylated residue- IJMS 2024

Polyomavirus Large T-antigen (LT) is the major viral regulatory protein that targets numerous cellular factors/pathways for cellular transformation and viral replication. LT directly recruits the cellular replication factors involved in recognition of the viral origin, origin unwinding, and primer synthesis carried out by mutual interactions between LT, DNA polymerase alpha-primase (Polprim), and single-stranded DNA binding complex, (RPA). The activities as well as protein-protein interactions of these complexes, are known to be modulated by post-translational modifications; however, modern high-sensitivity proteomic analyses of the PTMs as well as proteins associated with the three have been lacking. Using high resolution liquid chromatography tandem mass spectrometry (LC-MS/MS) of the immunoprecipitated factors (IPMS) we identified 602 PAARs for the 8 polypeptides of LT, Polprim and RPA, of which 479 had not previously been identified: 82 PAARs on SV40 LT, 305 on the Polprim heterotetrametric complex and 92 on the RPA heterotrimeric complex. Using phosphomimetic mutation of one of the novel phosphorylated aa residues, Threonine 518 detected on LT in this study, we have demonstrated a dramatic decrease in DNA replication functions of SV40 Large T- antigen both in vitro and in cell culture. The expansion of our understanding of PAAR sites provides new avenues to investigate regulation of important viral-host interactions with implications for devising targeted therapeutic strategies.
The MS data includes raw data as excel files from the UB Mass Spec Facility with Sequence maps in paper.
Excel documents: SV40 LT (Cont and Etoposide treated: ETO), POLA (Cont and ETO) and RPA (Cont and ETO).

Following are brief explanations of all columns in the tables:
Sequence - the peptide sequence identified
# PSMs - how many times a spectrum has been matched to this peptide sequence ( a crude quantitative value)
# Proteins - how many protein(s) this peptide can be inferred to
# Protein groups - similar with above but several proteins can be grouped to one protein group if no unique peptides can be identified to discriminated one from the other
Protein group accessions - Uniprot (or other protein database) accession number
Modifications - PTMs identified on the peptide
deltaCn - the difference of XCorr (search engine score) between the best match and the second best match (which is a match to another peptide, usually only the best match is considered as the "correct" match)
phosphoRS site probabilities - Predicted probabilities of PTM localization
XCorr - search engine score from Sequest HT, the higher XCorr the more confident the identification
Charge, MH+, RT - basic probabilities of the peptide ion
deltaM - mass difference between the theoretical and detected peptide ion
# Missed cleavages - how many K/Rs left in the peptide sequence, which should be cleaved by trypsin but are not