No modifications identified using open search with MS Fragger/Philosopher in PD 2.2

[kallepult]

Dear Huiyin,

With your highly appreciated help I have successfully run closed searches with MS Fragger/Philosopher (PeptideProphet) in PD 2.2 using the closed search template workflow, identifying various modifications including phosphorylation and oxidation on various amino acids. However, when trying the open search workflow with MS Fragger/Peptide Prophet, I am not able to identify any modifications from the same raw file (I do identify non-modified peptides), unless I specify them as variable modifications in the MS Fragger node.

The Philosopher node does give me some warnings as seen in the attached MagellanServer.log, but I'm not sure whether these are related to the issue.

Do you have any suggestions to what I might be doing wrong?

Thanks again for your help!

Best regards, Kasper

MagellanServer.log

[huiyinc]

Hi Kasper,

When you run open search without specifying any variable/fix modification in MSFragger node, it is expected to have no modification listed in the modification column of PSM table. This is because there is no predefined modification, but only the deltaM/Z values. Maybe you could use our FragPipe (which has more functionalities for open search) if you are looking for new/undiscovered modifications. Thanks.

Huiyin

kallepult notifications@github.com 於 2020年4月2日 週四 上午9:36寫道：

Dear Huiyin,

With your highly appreciated help I have successfully run closed searches with MS Fragger/Philosopher (PeptideProphet) in PD 2.2 using the closed search template workflow, identifying various modifications including phosphorylation and oxidation on various amino acids. However, when trying the open search workflow with MS Fragger/Peptide Prophet, I am not able to identify any modifications from the same raw file (I do identify non-modified peptides), unless I specify them as variable modifications in the MS Fragger node.

The Philosopher node does give me some warnings as seen in the attached MagellanServer.log, but I'm not sure whether these are related to the issue.

Do you have any suggestions to what I might be doing wrong?

Thanks again for your help!

Best regards, Kasper

MagellanServer.log https://github.com/Nesvilab/PD-Nodes/files/4421364/MagellanServer.log

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-- Hui-Yin Chang, 張彙音 Research Fellow Department of Pathology University of Michigan

[kallepult]

Dear Huiyin,

Thanks for your quick reply! If the open search option is not able to identify modifications that are not specified, what is the purpose of the open search workflow compared to the closed search?

I am attempting to identify (and quantify) peptides with a large number of different modifications (around 15), so I was hoping that an open search would give better results than closed ones with this many modifications normally do.

I will try out FragPipe instead - any chance that this functionality will make it into the PD nodes at one point?

Thanks again for your help!

Best regards, Kasper

[huiyinc]

Hi Kasper,

Yes, that's what we expect in open search. We are continually working on the improvement of open search in PD, but it takes time. That's the reason why I would suggest you to use FragPipe. Hope we will have more accurate open search results via MSFragger-PD node soon. Thanks.

Huiyin

kallepult notifications@github.com 於 2020年4月2日 週四 上午10:06寫道：

Dear Huiyin,

Thanks for your quick reply! If the open search option is not able to identify modifications that are not specified, what is the purpose of the open search workflow compared to the closed search?

I am attempting to identify (and quantify) peptides with a large number of different modifications (around 15), so I was hoping that an open search would give better results than closed ones with this many modifications normally do.

I will try out FragPipe instead - any chance that this functionality will make it into the PD nodes at one point?

Thanks again for your help!

Best regards, Kasper

— You are receiving this because you commented. Reply to this email directly, view it on GitHub https://github.com/Nesvilab/PD-Nodes/issues/5#issuecomment-607867942, or unsubscribe https://github.com/notifications/unsubscribe-auth/ALAWWA7ZLODEP2W55NUYL2TRKSLXXANCNFSM4L2OD7YQ .

-- Hui-Yin Chang, 張彙音 Research Fellow Department of Pathology University of Michigan

[anesvi]

Kasper,

Open search of course identifies modified peptides. But modifications are encoded just as mass shifts, e.g. if you look for phosphorylation using open search, It will not show up as MNCS[Phospho]DER peptide. Instead, it will be MNCSDER (but calculated-measured mass, delta mass value) will be ~ +80. I assume there is delta mass or mass diff column somewhere, that you can use to sort your data and look for modified peptides? So MSFragger is doing open search is intended. But PD may have limited functionality for showing open search results.

Best Alexey

From: chuiyin notifications@github.com Sent: Thursday, April 2, 2020 10:15 AM To: Nesvilab/PD-Nodes PD-Nodes@noreply.github.com Cc: Subscribed subscribed@noreply.github.com Subject: Re: [Nesvilab/PD-Nodes] No modifications identified using open search with MS Fragger/Philosopher in PD 2.2 (#5)

External Email - Use Caution Hi Kasper,

Yes, that's what we expect in open search. We are continually working on the improvement of open search in PD, but it takes time. That's the reason why I would suggest you to use FragPipe. Hope we will have more accurate open search results via MSFragger-PD node soon. Thanks.

Huiyin

kallepult notifications@github.com<mailto:notifications@github.com> 於 2020年4月2日 週四 上午10:06寫道：

Dear Huiyin,

Thanks for your quick reply! If the open search option is not able to identify modifications that are not specified, what is the purpose of the open search workflow compared to the closed search?

I am attempting to identify (and quantify) peptides with a large number of different modifications (around 15), so I was hoping that an open search would give better results than closed ones with this many modifications normally do.

I will try out FragPipe instead - any chance that this functionality will make it into the PD nodes at one point?

Thanks again for your help!

Best regards, Kasper

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-- Hui-Yin Chang, 張彙音 Research Fellow Department of Pathology University of Michigan

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[kallepult]

Dear Huiyin and Alexey,

Thanks again for your rapid replies and explanation! I will try out FragPipe and potentially also use the delta m/z values displayed in PD as well. Thank you for your great work and I hope to see the feature of matching delta m/z values with modifications in the PD node at some point!

Best regards, Kasper