Raw data were processed using Proteome Discoverer 2.2.0.388 (Thermo Fisher Scientific) and Mascot Server 2.6.2 (Matrix Science, London). A processing workflow was designed to search the MS2 data against the UniProt/SwissProt mouse protein database (07.2017 release, 16,909 entries) using trypsin/P as an enzyme, a maximum of two missed cleavage sites, 10 ppm and 0.5 Da, as the precursor, and fragment ion mass tolerances, respectively. Carbamidomethylated cysteines (+57.02146 Da), TMT10 labeled lysines and peptide N-termini (+229.162932 Da) were set as static, while oxidized methionines (+15.99492 Da) were set as dynamic modifications. The peptide-to-spectrum matches (PSMs) false discovery rates (FDRs) were controlled using Percolator and setting a max. delta Cn of 0.05 and a q-Value threshold of 0.01 (strict). Reporter ion quantitation was performed using the MS3 data order, 3 mmu peak integration and most confident centroid tolerances. Reporter ion intensities were adjusted to correct for the isotopic impurities of the
different TMT reagents (manufacturer specifications). A consensus workflow was defined to group PSMs into peptide and proteins. PSMs from all ranks were considered, peptide FDRs were controlled by setting a q-Value threshold of 0.01 and allowing the software to automatically select PSM q-Value or ion score for the grouping (PSM FDR was 1.0%, peptide group FDR was 1.8%). High confidence peptides with a minimal length of 6 residues were further grouped into proteins and protein FDR was set to fulfill a q-Value threshold of 0.01. At this level, protein FDR was 1.0%. For protein grouping, strict parsimony principle was applied. Peptide and protein quantitation was performed by summing intensities for each channel and normalizing each value with the highest TMT channel total. For protein quantitation, unique peptides only were considered. Finally, individual peptide and protein intensities were scaled to an average of 100.
[See
results attachment job
for details]
Keywords: N/A
Number of Files: | |
Total Size: | |
Experimental Design | |
Conditions:
|
|
Biological Replicates:
|
|
Technical Replicates:
|
|
Identification Results | |
Proteins (Human, Remapped):
|
|
Proteins (Reported):
|
|
Peptides:
|
|
Variant Peptides:
|
|
PSMs:
|
|
Quantification Results | |
Differential Proteins:
|
|
Quantified Proteins:
|
|
Browse Reanalysis Files | |
FTP Download Link (click to copy):
|