Data dependent acquisition (DDA) for raw spectral library
construction
Peak lists were generated from DDA raw data with DataAnalysis 4.4
(Bruker Daltonics) and peptide to spectrum matches identified with PEAKS
suite X plus (Bioinformatics Solutions Inc., Waterloo, Canada) and
X!Tandem Alanine (The GPM) database search engines followed by
unambiguous assignment of peptides to unique proteins using the O.
niloticus proteome database downloaded from NCBI RefSeq on Feb. 25,
2020. The database searched included 61,681 proteins plus the same
number of randomly scrambled decoys and 282 common contaminants (human
keratins, porcine trypsin, etc.). Trypsin cleavage was specified with at
the C-terminus of either Lys or Arg except when followed by Pro, and two
missed cleavages were the allowed maximum. Further, possible PTMs were
searched first by allowing for Cys carbamidomethylation, Met oxidation,
and Protein N-terminal acetylation. A second-round search considered all
313 PEAKSPTM modifications included in the default PEAKS suite X plus
database, allowing max. 3 PTMs per peptide. Mass tolerance limits were
set at 10 ppm for precursors and 0.03 Da for fragment ions. All DDA data
and metadata are available at MassIVE (MSV000085637,
doi:10.25345/C5DQ74) and ProteomeXchange (PXD020056).