/maxquant-proteomics

name: "maxquant-proteomics"
description: "MaxQuant + Perseus proteomics pipeline: run MaxQuant for LFQ and SILAC; parse proteinGroups.txt in Python; filter contaminants/decoys; log2 + median-normalize; impute MNAR; t-test with FDR; volcano plot; GO/pathway enrichment. Use Proteome Discoverer for Thermo-native processing; FragPipe/MSFragger for GPU-accelerated DB search."
license: "Apache-2.0"

MaxQuant + Perseus — Proteomics Analysis Pipeline

Overview

MaxQuant is the community-standard software for label-free quantification (LFQ) and SILAC proteomics. It performs database search, protein grouping, and intensity-based quantification from raw LC-MS/MS files, producing `proteinGroups.txt` as the primary output. Downstream statistical analysis — filtering, normalization, imputation, differential abundance testing, and visualization — is performed in Python using pandas, scipy, and matplotlib/seaborn, mirroring the Perseus workflow in a reproducible scripting environment.

When to Use

• Performing label-free quantification (LFQ) of proteins across multiple biological conditions — MaxQuant's MaxLFQ algorithm is the community benchmark
• Running SILAC (stable isotope labeling) experiments with light/heavy or triple-label designs
• Processing iTRAQ or TMT isobaric labeling experiments via MaxQuant's reporter ion quantification
• Identifying and quantifying proteins when you need the widely-cited MaxQuant output format (`proteinGroups.txt`) for comparison with published datasets
• Performing statistical differential abundance analysis on MaxQuant outputs without installing Perseus (GUI-only, Windows)
• Generating publication-quality volcano plots and GO enrichment from proteomics data in a reproducible Python workflow
• Use **Proteome Discoverer** instead when working with Thermo raw files requiring instrument-native processing or Sequest HT
• Use **FragPipe/MSFragger** instead for GPU-accelerated database search (3–10× faster) or when processing DIA (data-independent acquisition) data
• Use **omics-plotting** SKILL after differential abundance analysis or GSEA for publication-quality plots

Prerequisites

• **MaxQuant**: Windows software; download from https://maxquant.org/ (v2.4+); requires .NET 6 runtime
• **Python packages**: `pandas`, `numpy`, `scipy`, `matplotlib`, `seaborn`, `statsmodels`, `gseapy`
• **Data requirements**: Thermo `.raw` files or mzML-converted files; FASTA protein database (UniProt reviewed + contaminant database)
• **Environment**: MaxQuant runs on Windows (GUI or CLI); Python analysis runs cross-platform

pip install pandas numpy scipy matplotlib seaborn statsmodels gseapy

# Install pyMaxQuant for programmatic mqpar.xml configuration
pip install pymaxquant

Quick Start

import pandas as pd
import numpy as np

# Load MaxQuant output
df = pd.read_csv("combined/txt/proteinGroups.txt", sep="\t", low_memory=False)
print(f"Raw protein groups: {len(df)}")

# Filter contaminants, reverse decoys, only-by-site
mask = (
    (df["Potential contaminant"] != "+") &
    (df["Reverse"] != "+") &
    (df["Only identified by site"] != "+")
)
df = df[mask].copy()
print(f"After filtering: {len(df)} protein groups")

# Extract LFQ intensity columns
lfq_cols = [c for c in df.columns if c.startswith("LFQ intensity ")]
print(f"LFQ columns: {lfq_cols}")

# Log2-transform (0 → NaN)
lfq = df[lfq_cols].replace(0, np.nan)
lfq = np.log2(lfq)
print(f"Valid values per sample:\n{lfq.notna().sum()}")

Workflow

Step 1: Configure MaxQuant Parameters via mqpar.xml

MaxQuant is controlled by an XML parameter file (`mqpar.xml`). Edit it programmatically to set file paths, enzyme, modifications, and quantification type before running the search.

import xml.etree.ElementTree as ET

def update_mqpar(template_path: str, output_path: str,
                 raw_files: list[str], fasta_path: str,
                 experiment_names: list[str]) -> None:
    """Update mqpar.xml with sample-specific file paths."""
    tree = ET.parse(template_path)
    root = tree.getroot()

    # Set raw file paths
    file_paths_node = root.find(".//filePaths")
    file_paths_node.clear()
    for rf in raw_files:
        elem = ET.SubElement(file_paths_node, "string")
        elem.text = rf

    # Set experiment names (maps files to conditions)
    experiments_node = root.find(".//experiments")
    experiments_node.clear()
    for name in experiment_names:
        elem = ET.SubElement(experiments_node, "string")
        elem.text = name

    # Set FASTA database
    fasta_node = root.find(".//fastaFiles/FastaFileInfo/fastaFilePath")
    fasta_node.text = fasta_path

    tree.write(output_path, xml_declaration=True, encoding="utf-8")
    print(f"Written: {output_path}")

# Example usage
raw_files = [
    r"C:\Data\ctrl_rep1.raw",
    r"C:\Data\ctrl_rep2.raw",
    r"C:\Data\treat_rep1.raw",
    r"C:\Data\treat_rep2.raw",
]
update_mqpar(
    template_path="mqpar_template.xml",
    output_path="mqpar.xml",
    raw_files=raw_files,
    fasta_path=r"C:\Databases\human_uniprot_contaminants.fasta",
    experiment_names=["ctrl", "ctrl", "treat", "treat"],
)

Key `mqpar.xml` parameters (set in template or edit directly):

<!-- Enzyme and search settings -->
<enzymes>
  <string>Trypsin/P</string>
</enzymes>
<maxMissedCleavages>2</maxMissedCleavages>
<variableModifications>
  <string>Oxidation (M)</string>
  <string>Acetyl (Protein N-term)</string>
</variableModifications>
<fixedModifications>
  <string>Carbamidomethyl (C)</string>
</fixedModifications>

<!-- LFQ settings -->
<lfqMode>1</lfqMode>                     <!-- 1 = LFQ enabled -->
<lfqMinRatioCount>2</lfqMinRatioCount>   <!-- minimum peptides for LFQ -->
<matchBetweenRuns>True</matchBetweenRuns>

<!-- FDR thresholds -->
<peptideFdr>0.01</peptideFdr>
<proteinFdr>0.01</proteinFdr>

Step 2: Run MaxQuant from Command Line (Windows)

MaxQuant can be run headlessly from the Windows command prom