1. Notes

2. Result Statistics

Figure 1. False discovery rate (FDR) curve. X axis is the number of peptide-spectrum matches (PSM) being kept. Y axis is the corresponding FDR.

Figure 2. PSM score distribution. (a) Distribution of PEAKS peptide score; (b) Scatterplot of PEAKS peptide score versus precursor mass error.

(a)

(b)

Figure 3. De novo result validation. Distribution of residue local confidence: (a) Residues in de novo sequences validated by confident database peptide assignment; (b) Residues in "de novo only" sequences.

(a)

(b)

Table 1. Statistics of data. # of MS scans 6123 # of MS/MS scans 122442 Table 2. Result filtration parameters. Peptide -10lgP ≥15 Peptide Ascore ≥0 Protein -10lgP ≥20 Proteins unique peptides ≥0 De novo ALC Score ≥50% Table 3. Statistics of filtered result. Peptide-Spectrum Matches 36717 Peptide sequences 15887 Protein groups 3325 Proteins 3399 Proteins (#Unique Peptides) 1282 (>2); 286 (=2); 429 (=1); FDR (Peptide-Spectrum Matches) 0.5% FDR (Peptide Sequences) 0.9% FDR (Protein) 48.0% De Novo Only Spectra 24225

Table 4. PTM profile. Name ∆Mass Position #PSM -10lgP Area AScore Carbamidomethyl 57.02 C 4206 64.65 1.28E6 1000.00 Oxidation 15.99 M 983 52.74 2.29E5 1000.00

3. Experiment Control

Figure 4. Precursor mass error of peptide-spectrum matches (PSM) in filtered result. (a) Distribution of precursor mass error in ppm; (b) Scatterplot of precursor m/z versus precursor mass error in ppm.

(a)

(b)

Table 5. Number of identified peptides in each sample by the number of missed cleavages

Missed Cleavages 0 1 2 3 4+ Sample 2 14467 1331 77 12 0

4. Other Information

Table 6. Search parameters. Search Engine Name: PEAKS Parent Mass Error Tolerance: 0.07 Da Fragment Mass Error Tolerance: 0.025 Da Precursor Mass Search Type: monoisotopic Enzyme: Trypsin Max Missed Cleavages: 3 Non-specific Cleavage: both Fixed Modifications:   Carbamidomethylation: 57.02 Variable Modifications:   Oxidation (M): 15.99 Max Variable PTM Per Peptide: 3 Database: H.Sapiens030722 Taxon: All Searched Entry: 20384 FDR Estimation: Enabled Different data refine parameters are used for this search:

Table 7. Instrument parameters. Fractions: Lucena novo_1_RA2_01_1105.d, Lucena novo_2_RA2_01_11 06.d Ion Source: ESI(nano-spray) Fragmentation Mode: CID, CAD(y and b ions) MS Scan Mode: Quadrupole MS/MS Scan Mode: Time of Flight (TOF)