Fig 1.
Schematic workflow for the proteome analysis of (A) fetal bovine serum (FBS) and (B) cell secretome.
(A) FBS by itself as well as the residual FBS in the conditioned media or on the surface of the culture plate were analyzed. MS/MS data were compared against a bovine sequence database and the resultant protein list was used for the construction of an FBS database. (B) MS/MS data from cell secretome analysis were compared against various databases including human, human plus bovine and human plus FBS databases.
Fig 2.
Search results of MS/MS data from Calu-1 secretome analysis vary depending on the database used.
(A) PSMs are classified based on their species specificity. The classifications are PSMs belonging to: the top 30 FBS proteins or the proteins identified from residual FBS in the media or on the surface of the plate (Major in FBS), other FBS proteins (Minor in FBS), bovine proteins not identified by FBS analysis (Other bovine), and none of the species exclusively due to identical sequences (Orthologous peptides). (B) The number of identified proteins. Bovine proteins are subgrouped as in (A). (C) Venn diagram showing human proteins among the identified proteins.
Fig 3.
Distribution of posterior error probability (PEP) and Xcorr values for PSMs of Calu-1 secretome fractionated by OFFGEL fractionator.
(A) -logPEP values are compared between each pair of peptide-spectrum matches (PSMs). The region where |Δ(-logPEP)| < 0.7 between HuDB and HFDB search results is marked grey. Spectra in blue area have PSMs in the HFDB, but not in HuDB search result. Spectra in red area have higher -logPEP value (more than 0.7) for the PSMs of HFDB than for HuDB search results. The reverse is true for the spectra in green area. (B) Distribution of XCorr values for HFDB vs. HuDB search results.
Fig 4.
Search results of MS/MS data from SILAC-labeled secretome of HCT-116.
(A) Mass spectrometric data were searched against HuDB, HBDB and HFDB databases, and then arginine, lysine, and proline residues in all PSMs were counted and classified into isotope-labeled (red) and unlabeled (blue) residues. (B) Pie-charts showing the number of proteins classified as human and bovine. Labeled: identification from all labeled peptides; partially labeled: identification from both labeled and unlabeled peptides; non-labeled: identification from unlabeled peptides only.