Table 4.
Parameters used for database-searching of LC-MS3 raw data of TMT-labeled peptides in Proteome Discoverer 2.4
| Parameter | Value |
|---|---|
| Modifications | |
| Fixed modifications | Carbamidomethyl (Cysteine) |
| TMT 10-Plex (Lysine, Peptide N-Terminus) | |
| Dynamic modifications | Oxidation (Methionine) |
| Acetyl (Protein N-Terminus) | |
| Deamidation (Protein N-Terminus) | |
| Methionine Loss (Protein-N-Terminus) | |
| Methionine Loss (Protein-N-Terminus) + Acetyl (Protein N-Terminus) | |
| PyroGlu (Glutamine) | |
| Digestion | |
| Protease | Trypsin (Proteolysis before Praline enabled) |
| Missed cleavages | 2 allowed |
| Peptide properties | |
| Maximum charge | 6 |
| Maximum amino acid number | 6 |
| Maximum amino acid number | 144 |
| Maximum weight (Da) | 350 |
| Minimum weight (Da) | 5000 |
| Mass tolerance | |
| Precursor mass tolerance (ppm) | 10 |
| Fragment mass tolerance (Da) | 0.6 |
| Reporter mass tolerance (ppm) | 20 |
| FDR calculation | |
| PSM FDR | < 0.01 |
| Peptide FDR | < 0.01 |
| Protein FDR | < 0.01 |
| Reporter quantification | |
| Peptides to use | Unique and Razor |
| Co-Isolation threshold | 50 |
| Average reporter Signal to noise ratio | 10 |
| SPS mass matches (%) | 65 |