Proteomic computational analysis

The raw data files were processed and quantified using PatternLab for Proteomics software [43] (version 4.0). Peptide sequence matching (PSM) was performed using the Comet algorithm against the protein-centric human database neXtProt [44] plus the SARS-CoV-2 reference proteome from UniProt [45] under ID UP000464024, both downloaded March 29, 2021. A target-decoy strategy was employed. The search parameters were tryptic and semitryptic peptides, with masses between 500 and 5000 Da, up to 2 lost cleavage sites; modifications: carbamidomethylation (Cys), oxidation (Met), and initial tolerance of 40 ppm for precursor ions. PSMs were filtered using the Search Engine Processor (SEPro) module, and identifications were grouped by the number of enzymatically cleaved ends, resulting in two distinct subgroups. For each result, the scores for each metric (XCorr, DeltaCN, and ZScore) were used to generate a Bayesian discriminator, accepting up to a 1% false discovery rate (FDR), estimated by the number of decoy sequence IDs. The results were further filtered to accept only PSMs with a mass error less than 5 ppm and protein identifications supported by two or more independent identifications. Proteins identified by a single spectrum (1 hit wonder) with XCorr below 2 were excluded. The final list of identified peptides and mapped proteins for all samples was reported. The list of resulting peptides from shotgun proteomics was used for alignment with the sequences of human endogenous retrovirus K113 (https://www.ncbi.nlm.nih.gov/nuccore/{"type":"entrez-nucleotide","attrs":{"text":"NC_022518.1","term_id":"548558394"}}NC_022518.1). The alignment was carried out with the NCBI/BLAST database through the Protein Blast — BlastP algorithm. Alignments with identity and coverage equal to or greater than 50% were considered. Detailed information about the proteins that aligned with the peptides can be obtained from the UniProtKB SwissProt database (https://www.uniprot.org/uniprot/?query=Human+endogenous+retrovirus+K113&sort=score).