multiFLEX-LF: A Computational Approach to Quantify the Modification Stoichiometries in Label-Free Proteomics Data Sets

J Proteome Res. 2022 Apr 1;21(4):899-909. doi: 10.1021/acs.jproteome.1c00669. Epub 2022 Jan 27.

Abstract

In liquid-chromatography-tandem-mass-spectrometry-based proteomics, information about the presence and stoichiometry of protein modifications is not readily available. To overcome this problem, we developed multiFLEX-LF, a computational tool that builds upon FLEXIQuant, which detects modified peptide precursors and quantifies their modification extent by monitoring the differences between observed and expected intensities of the unmodified precursors. multiFLEX-LF relies on robust linear regression to calculate the modification extent of a given precursor relative to a within-study reference. multiFLEX-LF can analyze entire label-free discovery proteomics data sets in a precursor-centric manner without preselecting a protein of interest. To analyze modification dynamics and coregulated modifications, we hierarchically clustered the precursors of all proteins based on their computed relative modification scores. We applied multiFLEX-LF to a data-independent-acquisition-based data set acquired using the anaphase-promoting complex/cyclosome (APC/C) isolated at various time points during mitosis. The clustering of the precursors allows for identifying varying modification dynamics and ordering the modification events. Overall, multiFLEX-LF enables the fast identification of potentially differentially modified peptide precursors and the quantification of their differential modification extent in large data sets using a personal computer. Additionally, multiFLEX-LF can drive the large-scale investigation of the modification dynamics of peptide precursors in time-series and case-control studies. multiFLEX-LF is available at https://gitlab.com/SteenOmicsLab/multiflex-lf.

Keywords: LC−MS/MS; PTM quantification; bioinformatics tool; label-free quantification; modification stoichiometry; post-translational modification.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Chromatography, Liquid
  • Mass Spectrometry
  • Peptides
  • Proteins*
  • Proteomics*

Substances

  • Peptides
  • Proteins