Labelfree Protein Quantification

Disclaimer this tool needs a quantAndProt file, which combines the results from ProteinInference and PSMBasedQuantification.

After quantification and protein inference are performed, it is known which peptide originated from which protein, as well as the intensity of each peptide. The information available for each peptide now needs to be aggragated for their proteins.

This tool performs the aggregation from the peptides to the protein in several steps. The first two aggregation steps are optional. One of them is the aggregation based on charge state. Peptides with the same sequence and modifications, but different charge states are being aggregated. The next optional step does the same for peptides with the same sequence, but different modifications. Those steps build upon each other. The last step is the aggregation of all peptides of a protein. The result of each aggregation step is given as a tab separated file. The aggregation is performed according to the given parameters for each step. If an optional aggregation is not performed, the next step takes the result from the prior aggregation. For example, if aggregation by charge and modification are skipped, the protein aggregation is performed on previously unaggregated peptides, where a peptidesequence can occur with different charge states and modifications.

Parameters

LabelFreeQuantificationParams

The following table gives an overview of the parameter set:

LabelFreeQuantification.AggregationParams

The following table gives an overview of the parameter set:

Parameter Generation

Parameters are handed to the cli tool as a .json file. you can download the default file here: minimal, with charge aggregation, with charge and modification aggregation. Alternatively, you can use an F# script, which can be downloaded or run in Binder at the top of the page, to write your own parameter file:

#r "nuget: ProteomIQon, 0.0.7"

open ProteomIQon
open ProteomIQon.Dto

let defaultLabelFreeQuantificationParams :Dto.LabelFreeQuantificationParams = 
    let chargeParams :Common.LabelFreeQuantification.AggregationParams = 
        {
                Transform        = None
                SingleFilters    = None
                GroupFilters     = None
                Aggregation      = {Light= NumericAggregation.Mean}
        }
    let modParams :Common.LabelFreeQuantification.AggregationParams = 
        {
                Transform        = None
                SingleFilters    = None
                GroupFilters     = None
                Aggregation      = {Light= NumericAggregation.Mean}
        }
    let protParams :Common.LabelFreeQuantification.AggregationParams = 
        {
                Transform        = None
                SingleFilters    = None
                GroupFilters     = None
                Aggregation      = {Light= NumericAggregation.Mean}
        }
    {
        ModificationFilter                   = UseModifiedPeptides.All 
        AggregatePeptideChargeStatesParams   = Some modParams // or None
        AggregateModifiedPeptidesParams      = Some modParams // or None
        AggregateToProteinGroupsParams       = protParams
    }   

let serialized = 
    defaultLabelFreeQuantificationParams
    |> Json.serialize

Executing the Tool

Disclaimer this tool needs a quantAndProt file, which combines the results from ProteinInference and PSMBasedQuantification.

To generate a quantified protein output for your run simply call:

proteomiqon-labelfreeproteinquantification -i "path/to/your/run/quantAndProt" -o "path/to/your/outDirectory" -p "path/to/your/params.json"

It is also possible to call the tool on a lists of quantAndProt files:

proteomiqon-labelfreeproteinquantification -i "path/to/your/run1.quantAndProt" "path/to/your/run2.quantAndProt" "path/to/your/run3.quantAndProt" "path/to/your/outDirectory" -p "path/to/your/params.json" 

A detailed description of the CLI arguments the tool expects can be obtained by calling the tool:

proteomiqon-labelfreeproteinquantification --help