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vdemichev / DiaNN

DIA-NN - a universal automated software suite for DIA proteomics data analysis.
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same peptide seq but different STY localization probability of same Serine amino acid #1234

Open singhsk2622 opened 3 weeks ago

singhsk2622 commented 3 weeks ago

dia_nn_sty Hi, I did phospho analysis in Fragpipe-DIA-NN. I see one strange thing in output file. I see that for same protein there 3 STY peptides has been identified. All three peptides have same sequence only difference in localization probability of same Serine amino acid. why it is assigning three different peptides and assigning in different rows? Any one explain it me?

vdemichev commented 3 weeks ago

Hi,

Can you please paste here the whole rows you are showing along with column names?

Best, Vadim

singhsk2622 commented 3 weeks ago

<html xmlns:v="urn:schemas-microsoft-com:vml" xmlns:o="urn:schemas-microsoft-com:office:office" xmlns:x="urn:schemas-microsoft-com:office:excel" xmlns="http://www.w3.org/TR/REC-html40">

Protein.Group | Protein.Ids | Protein.Names | Genes | First.Protein.Description | Proteotypic | Stripped.Sequence | Modified.Sequence | Precursor.Charge | Precursor.Id | E:\DATA\Mette_sty_4343-4375\H2_ANP_Slot2-33_1_4351.d | E:\DATA\Mette_sty_4343-4375\H1_CTRL_Slot2-28_1_4341.d | E:\DATA\Mette_sty_4343-4375\H5_CNP_Slot2-41_1_4367.d | E:\DATA\Mette_sty_4343-4375\H6_CNP_Slot2-44_1_4373.d | E:\DATA\Mette_sty_4343-4375\H1_CNP_Slot2-29_1_4343.d | E:\DATA\Mette_sty_4343-4375\H6_ANP_Slot2-45_1_4375.d | E:\DATA\Mette_sty_4343-4375\H2_CNP_Slot2-32_1_4349.d | E:\DATA\Mette_sty_4343-4375\H5_ANP_Slot2-42_1_4369.d | E:\DATA\Mette_sty_4343-4375\H11_ANP_Slot2-30_1_4345.d | E:\DATA\Mette_sty_4343-4375\H2_CTRL_Slot2-31_1_4347.d | E:\DATA\Mette_sty_4343-4375\H4_ANP_Slot2-39_1_4363.d | E:\DATA\Mette_sty_4343-4375\H5_CTRL_Slot2-40_1_4365.d | E:\DATA\Mette_sty_4343-4375\H3_CTRL_Slot2-34_1_4353.d | E:\DATA\Mette_sty_4343-4375\H3_ANP_Slot2-36_1_4357.d | E:\DATA\Mette_sty_4343-4375\H6_CTRL_Slot2-43_1_4371.d | E:\DATA\Mette_sty_4343-4375\H4_CNP_Slot2-38_1_4361.d | E:\DATA\Mette_sty_4343-4375\H4_CTRL_Slot2-37_1_4359.d | E:\DATA\Mette_sty_4343-4375\H3_CNP_Slot2-35_1_4355.d | All Mapped Proteins | All Mapped Genes | M:15.9949 | M:15.9949 Best Localization | M:15.9949 Best Scan | STY:79.96633 | STY:79.96633 Best Localization | STY:79.96633 Best Scan | n:42.0106 | n:42.0106 Best Localization | n:42.0106 Best Scan -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- | -- F1M6Z4 | F1M6Z4 |   | Prob1 |   | 1 | AASAPPTDRLHPASSLGMR | AAS(UniMod:21)APPTDRLHPASSLGM(UniMod:35)R | 2 | AAS(UniMod:21)APPTDRLHPASSLGM(UniMod:35)R2 | 4743.61 | 1802.44 | 620.495 | 1679.9 | 3714.83 | 2351.35 | 2216.74 | 1634.64 | 4171.05 | 2327.22 | 1572.71 | 593.243 | 616.555 | 2498.75 |   | 2767.73 | 1533.18 |   | tr\|F1M6Z4\|F1M6Z4_RAT | Prob1 | AASAPPTDRLHPASSLGM(1.0000)R | 1 | H1_CNP_Slot2-29_1_4343_diatracer.815118.815118.2 | AAS(0.8971)APPT(0.0277)DRLHPAS(0.0379)S(0.0373)LGMR | 0.8971 | H1_CNP_Slot2-29_1_4343_diatracer.815118.815118.2 F1M6Z4 | F1M6Z4 |   | Prob1 |   | 1 | AASAPPTDRLHPASSLGMR | AAS(UniMod:21)APPTDRLHPASSLGM(UniMod:35)R | 3 | AAS(UniMod:21)APPTDRLHPASSLGM(UniMod:35)R3 | 172206 | 54009.6 | 53949.4 | 99303.2 | 148441 | 120861 | 118466 | 78399.5 | 165968 | 118161 | 93134.4 | 72153.6 | 52809.7 | 103080 | 81127.9 | 127826 | 70269.4 | 64540.8 | tr\|F1M6Z4\|F1M6Z4_RAT | Prob1 | AASAPPTDRLHPASSLGM(1.0000)R | 1 | H11_ANP_Slot2-30_1_4345_diatracer.1040581.1040581.3 | AAS(0.9302)APPT(0.0222)DRLHPAS(0.0239)S(0.0238)LGMR | 0.9302 | H4_CTRL_Slot2-37_1_4359_diatracer.782931.782931.3 F1M6Z4 | F1M6Z4 |   | Prob1 |   | 1 | AASAPPTDRLHPASSLGMR | AAS(UniMod:21)APPTDRLHPASSLGM(UniMod:35)R | 4 | AAS(UniMod:21)APPTDRLHPASSLGM(UniMod:35)R4 | 3602.33 |   |   | 8367.68 | 3951.48 | 6857.46 |   | 5254.43 | 3482.65 |   | 3302.2 |   |   |   |   |   |   |   | tr\|F1M6Z4\|F1M6Z4_RAT | Prob1 | AASAPPTDRLHPASSLGM(1.0000)R | 1 | H5_ANP_Slot2-42_1_4369_diatracer.447942.447942.4 | AAS(0.7913)APPT(0.0971)DRLHPAS(0.0391)S(0.0725)LGMR | 0.7913 | H5_ANP_Slot2-42_1_4369_diatracer.447942.447942.4

vdemichev commented 3 weeks ago

Charge states are different

singhsk2622 commented 3 weeks ago

Should not be merged in final file? now when i am doing statistical analysis so i will have 3 different fold change for three same peptides with different charge. Any suggestion for this ?

vdemichev commented 3 weeks ago

You can either aggregate yourself based on the main report or (if using DIA-NN 1.9 or later) use the phosphosite_90/99 matrices, how these are calculated is described here https://github.com/vdemichev/DiaNN?tab=readme-ov-file#ptms-and-peptidoforms.

singhsk2622 commented 3 weeks ago

thanks