PSMBasedQuantificationTIMs

PSMBasedQuantificationTIMs is the PSMBasedQuantification variant for timsTOF runs. On these instruments a peptide ion is a peak in two dimensions, retention time and ion mobility, so the tool extracts an intensity grid over that plane around every identification instead of a one dimensional ion chromatogram. By default it locates the peak on the grid with a Gabor filter (a Gaussian envelope multiplied with a cosine wave, tuned in size, width, frequency and angle), then fits and integrates it. The output has the same layout as the output of PSMBasedQuantification, so the downstream tools do not care which of the two produced it.

Inputs and outputs

The tool refuses any instrument file whose name does not end in .mzlite. It always reads the qpsm files as the FragPipe layout written by MsFraggerToPSM (with Hyperscore and IonMobility columns), so the files from PSMStatistics do not fit here. With one file per flag the pair is used as given. With lists, the n-th mzlite file is paired with the n-th qpsm file unless you pass -mf, which pairs files by their base name and skips mzlite files without a partner.

For every run the tool writes run.quant, tab separated with a header and the columns described on the PSMBasedQuantification page. QuantBasedAlignment, AlignmentBasedQuantification, AlignmentBasedQuantStatistics and JoinQuantPepIonsWithProteins read .quant files.

With -dc the tool writes charts into run_plots, and -z packs that directory into run_plots.zip and deletes it. Delete an existing run.quant in the output directory before a rerun, the tool appends to it. Logs go to PSMBasedQuantification_log.txt and run_log.txt in the output directory.

Parameters

The parameter record is the one of PSMBasedQuantification. The defaults below are the timsTOF defaults.

The Gabor parameters. The tool rejects a kernel size at or below zero and any width, frequency or angle that is not finite (widths must also be positive).

The default file is QuantificationTIMsParams.json.

Writing a parameter file

open ProteomIQon
open ProteomIQon.Domain

let gaborParams : Gabor3DParams =
    {
        sizeX     = 41
        sizeY     = 41
        sigmaX    = 8.0
        sigmaY    = 8.0
        frequency = 2.0
        theta     = 0.06
    }

let quantificationTIMsParams : Dto.QuantificationParams =
    {
        PerformLabeledQuantification = Labeling.Unlabeled
        FragPipe                     = true
        XicExtraction =
            {
                ScanTimeWindow = 2.
                MzWindow_Da    = Window.Estimate
                XicProcessing  = XicProcessing.Gabor3D gaborParams
                TopKPSMs       = None
            }
        BaseLineCorrection = None
    }

// Replace the temp folder with your project folder.
let outputPath = System.IO.Path.Combine(System.IO.Path.GetTempPath(), "QuantificationTIMsParams.json")

Json.serializeAndWrite outputPath quantificationTIMsParams

Running the tool

Install with dotnet tool install --global ProteomIQon.PSMBasedQuantificationTIMs, then quantify one run:

proteomiqon-psmbasedquantificationtims -i path/to/run.mzlite -ii path/to/run.qpsm -d path/to/database.db -o path/to/output -p path/to/QuantificationTIMsParams.json

Several runs, paired by position in the two lists, three in parallel:

proteomiqon-psmbasedquantificationtims -i path/to/run1.mzlite path/to/run2.mzlite path/to/run3.mzlite -ii path/to/run1.qpsm path/to/run2.qpsm path/to/run3.qpsm -d path/to/database.db -o path/to/output -p path/to/QuantificationTIMsParams.json -c 3

Two directories, paired by file name:

proteomiqon-psmbasedquantificationtims -i path/to/mzliteFolder -ii path/to/qpsmFolder -d path/to/database.db -o path/to/output -p path/to/QuantificationTIMsParams.json -c 3 -mf

All flags:

proteomiqon-psmbasedquantificationtims --help