Table I Relative quantification of phosphopeptides

MS-based relative quantification of tryptic SLP-65 phosphopeptides obtained from DT40 B cells, which were left untreated or BCR-stimulated for 2 or 20 minutes. The table lists the sequence, molecular weight (MW) and charge state (n) of the phosphopeptides, the MASCOT peptide score, the number of sites (no.), their exact positions (site), the number of biological replicates (R), the quantitative ratio of 0 versus 2 min (0′/2′) and 0 versus 20 min (0′/20′) as determined by MSQuant, and the standard deviation(s) (STD). Those values, which were calculated from peptides that could be quantified with only one biological sample, are marked with an asterisk. In those cases STD refers to values that were obtained solely by the number of MS scans used for relative quantification in the MSQuant program. A double-asterisk represents the mean value of the STD of two experiments. Phosphosites that were unambiguously identified by manual inspection of the MS/MS spectra are shown in bold black italic capitals (pS/pT/pY). Peptides with one or two phosphorylation sites that could not be identified unambiguously are shown in bold black italic lower-case letters (s/t/y). Table with all sequences and values used for calculation and the manually annotated MS/MS spectra of the quantified peptides are shown in the supplemental data.

PeptideMW (+n)ScoreSiteNo.R0′/2′STD 0′/2′0′/20′STD 0′/20′
TGKDpTWDR1057,4681 (+2)24Thr-54112.6*0.1*1.6*0.4*
KKPPPpSLPR1098,6314 (+3)38Ser-65132.
IPSpSFPISR1082,5360 (+2)35Ser-130134.
KIPSSFPISRGEpYADNR2016,0122 (+3)14Tyr-1381112.9*1.6*5.5*1.0*
KPSLPpSPAAKPK1299,6942 (+3)53Ser-173150.
KPpSLPpSPAAKPK1379,6617 (+2)42Ser-170, Ser-173261.
ECamSDDEDNYIVPVDNDDDNpYIEPTESSTPPPAKPPVNR4382,8776 (+3)45Tyr-205119.2*3.5*4.5*1.9*
ECampSDDEDNpYIVPVDNDDDNpYIEPTESSTPPPAKPPVNR4542,8170 (+3)48Ser-188, Tyr-194, Tyr-2053110.1*2.3*
ECampSDDEDNYIVPVDNDDDNYIEPtEsstPPPAKPPVNR4462,7997 (+3)27Ser-188, Thr-209/Ser-211/Ser-212/Thr-213210.9*0.1*1.3*0.2*
ECcamSDDEDNYIVPVDNDDDNYIEPTESSpTPPPAKPPVNR4382,8390 (+3)52Thr-213111.7*0.2*2.3*0.2*
SALPTPPKPSLASDMQEVpYEVPEEEEELSPPPVTR3927,8283 (+3)42Tyr-249110.9*0.2*1.8*0.4*
SALPpTPPKPSLASDMoxQEVYEVPEEEEELpSPPPVTR4023,7866 (+3)25Thr-235, Ser-259211.0*0.1*0.9*0.1*
AQNAEHSHMoxHpSMTRESPK2188,8773 (+4)21Ser-285120.4**0.04**0.3**0.05**
EpSPKLDASR1081,4620 (+2)49Ser-290150.
FPPGAAPpSPLPR1285,6433 (+2)54Ser-339141.
RGpSSHEFPLPPLPSGTPK1983,0084 (+3)42Ser-381133.
RGpSSHEFPLPPLPSGpTPK2062,9731 (+3)32Ser-381, Thr-394223.30.7**1.80.4**
ssLQKPLVLPK1288,7360 (+2)23Ser-397/Ser-398134.70.67.14
ALGTpSPHSSISSISSTADQDAGVHSK2621,1562 (+2)85Ser-419121.
ALGTsPHssISSISSTADQDAGVHSK2701,1458 (+3)62Ser-419/Ser-422/Ser-423241*0.2*1.6*0.3*
TAEDALpYR1021,4243 (+2)44Tyr-457115.0*0.7*6.3*1.7*
KSpSGQDSR943,3749 (+2)22Ser-471111.2*0.2*6.1*0.83*