Fig. 4

Role of PhosphOrtholog in the MS-based PTM data analysis pipeline. Illustration of the broadly divided four stages of MS-based PTM experiments, in Stage 1, sample extraction and preparation tasks are conducted from human and rat muscle tissues for the MS-based phosphoproteomics experiment. Stage 2 marks the raw spectral data analysis to generate peptide and protein annotations along with intensity measures for the PTMs induced by the experimental design in each species. In Stage 3, output from Stage 2 is parsed to extract information such as the leading Uniprot ID (‘Uniprot_ACC’), modified amino acid type (‘AminoAcid’) and modification site number (‘Site#’) from each species and concatenated in the desired input format for PhosphOrtholog mapping. We showcase the PTM examples for proteins ULK1 (2 sites in human and rat) and ACACA (3 sites in human and rat) here; column ‘ModificationSite’ indicates the peptide sequence with the identified PTM site and the probability of particular amino acids being phosphorylated by the number within the parenthesis. In Stage 4, the sites mapped by PhosphOrtholog are obtained, which are either annotated as newly mapped with a calculated E-value (4 out of 5 input sites were not mapped before, identified with E-value of 0) or with “From PhosphoSitePlus” if the mapping was previously known (mapping between human ACACA site S80 and rat ACACA site S79 is annotated in PhosphoSitePlus database)