Plasma Precipitation Analysis
Created: 2012-07-31 16:04:54
Last updated: 2012-07-31 16:16:40
ABOUT THE WORKFLOW This workflow was used to analyze the data in a manuscript by Mostovenko et al. (2012, submitted), comparing proteins in the precipitate with those left in solution after organic solvent precipitation. However, the workflow is generally applicable in comparisons of any binary fractionation method in proteomics, where the fractions are analyzed by liquid chromatography-tandem mass spectrometry. The workflow identifies proteins by SpectraST spectral library search and X!Tandem database search, validates the results using PeptideProphet and combines them with InterProphet. The proteins are quantified by spectral counting and the fractions compared with respect to protein molecular weight, isoelectric point and GRAVY hydrophobicity score. In the end, an Rshell script produces the figures as presented in the manuscript. Executing the workflow requires Rserve running and the Trans-Proteomic Pipeline (http://tools.proteomecenter.org/wiki/index.php?title=Software:TPP) installed with default settings and in the default location. The Rshell script contains the location where the figures will be generated. All other output files are stored in the input data folders by default. The version of X!Tandem called from this workflow is separate from the one installed with the Trans-Proteomic Pipeline (the location specified in the Tandem components). COPYRIGHT NOTICE This workflow is based on open software and the workflow itself is shared under the Creative Commons Attribution-ShareAlike 3.0 Unported License (CC BY-SA) (see http://creativecommons.org/licenses/by-sa/3.0/) Copyright© 2012 Ekaterina Mostovenko, Jeroen de Bruin, Yassene Mohammed and Magnus Palmblad
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Authors (1)
| Ekaterina Mostovenko (e.mostovenko@lumc.nl)
Jeroen de Bruin
Yassene Mohammed
Magnus Palmblad (n.m.palmblad@lumc.nl) |
Titles (1)
| Plasma Precipitation Analysis V0.9 |
Descriptions (1)
| ABOUT THE WORKFLOW
This workflow was used to analyze the data in a manuscript by Mostovenko et al. (2012, submitted), comparing proteins in the precipitate with those left in solution after organic solvent precipitation. However, the workflow is generally applicable in comparisons of any binary fractionation method in proteomics, where the fractions are analyzed by liquid chromatography-tandem mass spectrometry.
The workflow identifies proteins by SpectraST spectral library search and X!Tandem database search, validates the results using PeptideProphet and combines them with InterProphet. The proteins are quantified by spectral counting and the fractions compared with respect to protein molecular weight, isoelectric point and GRAVY hydrophobicity score. In the end, an Rshell script produces the figures as presented in the manuscript.
Executing the workflow requires Rserve running and the Trans-Proteomic Pipeline (http://tools.proteomecenter.org/wiki/index.php?title=Software:TPP) installed with default settings and in the default location. The Rshell script contains the location where the figures will be generated. All other output files are stored in the input data folders by default. The version of X!Tandem called from this workflow is separate from the one installed with the Trans-Proteomic Pipeline (the location specified in the Tandem components).
COPYRIGHT NOTICE
This workflow is based on open software and the workflow itself is shared under the Creative Commons Attribution-ShareAlike 3.0 Unported License (CC BY-SA) (see http://creativecommons.org/licenses/by-sa/3.0/)
Copyright© 2012 Ekaterina Mostovenko, Jeroen de Bruin, Yassene Mohammed and Magnus Palmblad
|
Dependencies (0)
Inputs (6)
| Name |
Description |
| FASTA_file |
|
| Tandem_param_file |
|
| pellet_directory |
|
| spectral_library |
|
| raw_directory |
|
| supernatant_directory |
|
Processors (7)
| Name |
Type |
Description |
| compare_results |
workflow |
This embedded workflow calculates the molecular weight, isoelectric point and GRAVY score for each protein in the FASTA file and matches it with the protein identifications from the preceeding embedded workflows. An Rshell generates a Venn diagram for the identified peptides in the different fractions and histograms of protein spectral counts as function of the calculated protein properties. |
| analyze_raw |
workflow |
The analyze_raw subworkflow identifies proteins by SpectraST spectral library search and X!Tandem database search, validates the results using PeptideProphet and combines them with InterProphet. The PeptideProphet probability cut-off is specified the PeptideProphet tools, here 0.95. This probability cut-off typically gives a global false discovery rate around 1%. |
| analyze_pellet |
workflow |
The analyze_pellet subworkflow identifies proteins by SpectraST spectral library search and X!Tandem database search, validates the results using PeptideProphet and combines them with InterProphet. The PeptideProphet probability cut-off is specified the PeptideProphet tools, here 0.95. This probability cut-off typically gives a global false discovery rate around 1%. |
| analyze_supernatant |
workflow |
The analyze_supernatant subworkflow identifies proteins by SpectraST spectral library search and X!Tandem database search, validates the results using PeptideProphet and combines them with InterProphet. The PeptideProphet probability cut-off is specified the PeptideProphet tools, here 0.95. This probability cut-off typically gives a global false discovery rate around 1%. |
| CompassXport_raw |
workflow |
This embedded component converts liquid chromatography-tandem mass spectrometry data in the Bruker instrument format to the open mzXML format. This component should be replaced for data from other vendors. |
| CompassXport_pellet |
workflow |
This embedded component converts liquid chromatography-tandem mass spectrometry data in the Bruker instrument format to the open mzXML format. This component should be replaced for data from other vendors. |
| CompassXport_supernatant |
workflow |
This embedded component converts liquid chromatography-tandem mass spectrometry data in the Bruker instrument format to the open mzXML format. This component should be replaced for data from other vendors. |
Beanshells (14)
| Name |
Description |
Inputs |
Outputs |
| CompassXport |
|
input_directory
|
mzXML_file
|
| SpectraST |
|
library_splib_file
mzxml_file
parameters
|
output_file
|
| Tandem |
|
mzxml_file
fasta_file
parameter_file
|
tandem_file
|
| generate_InterProphet_filename |
|
tandem_filename
|
iproph_filename
|
| generate_InterProphet_filename |
|
tandem_filename
|
iproph_filename
|
| SpectraST |
|
library_splib_file
mzxml_file
parameters
|
output_file
|
| Tandem |
|
mzxml_file
fasta_file
parameter_file
|
tandem_file
|
| generate_InterProphet_filename |
|
tandem_filename
|
iproph_filename
|
| SpectraST |
|
library_splib_file
mzxml_file
parameters
|
output_file
|
| Tandem |
|
mzxml_file
fasta_file
parameter_file
|
tandem_file
|
| combine_IDs_and_properties_raw |
|
prot_list
ID
MW
Gravy_score
Prot_seq
pI
|
out_raw
|
| combine_IDs_and_properties_pellet |
|
prot_list
ID
MW
Gravy_score
Prot_seq
pI
|
out_pel
|
| combine_IDs_and_properties_supernatant |
|
prot_list
ID
MW
Gravy_score
Prot_seq
pI
|
out_sup
|
| get_properties |
|
seq
|
ID
Prot_seq
MW
Gravy_score
pI
|
Outputs (2)
| Name |
Description |
| Venn_diagram |
|
| histograms |
|
Datalinks (21)
| Source |
Sink |
| analyze_raw:InterProphet_filename |
compare_results:raw_input |
| analyze_pellet:InterProphet_filename |
compare_results:pellet_input |
| analyze_supernatant:InterProphet_filename |
compare_results:supernatant_input |
| FASTA_file |
compare_results:FASTA_file |
| FASTA_file |
analyze_raw:FASTA_file |
| Tandem_param_file |
analyze_raw:Tandem_param_file |
| spectral_library |
analyze_raw:spectral_library |
| CompassXport_raw:mzXML_file |
analyze_raw:mzXML_file |
| Tandem_param_file |
analyze_pellet:Tandem_param_file |
| spectral_library |
analyze_pellet:spectral_library |
| FASTA_file |
analyze_pellet:FASTA_file |
| CompassXport_pellet:mzXML_file |
analyze_pellet:mzXML_file |
| spectral_library |
analyze_supernatant:spectral_library |
| Tandem_param_file |
analyze_supernatant:Tandem_param_file |
| FASTA_file |
analyze_supernatant:FASTA_file |
| CompassXport_supernatant:mzXML_file |
analyze_supernatant:mzXML_file |
| raw_directory |
CompassXport_raw:input_directory |
| pellet_directory |
CompassXport_pellet:input_directory |
| supernatant_directory |
CompassXport_supernatant:input_directory |
| compare_results:Venn_diagram |
Venn_diagram |
| compare_results:histograms |
histograms |
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This is a rather large workflow, but it is a first attempt to encode in a single workflow all data analysis steps, from raw mass spectrometry data to figures as they appear in the paper. The database and spectral library searches are done in parallel in three identical embedded workflows rather than serially in one embedded workflow. This allows modification of or comparison between peptide identification strategies as well.
The Rshell script requires a package for the Venn diagram generation and the workflow assumes the Trans-Proteomic Pipeline is already installed, in its default location.
~Magnus