pepFunk: a tool for peptide-centric functional analysis of metaproteomic human gut microbiome studies

Bioinformatics. 2020 Aug 15;36(14):4171-4179. doi: 10.1093/bioinformatics/btaa289.


Motivation: Enzymatic digestion of proteins before mass spectrometry analysis is a key process in metaproteomic workflows. Canonical metaproteomic data processing pipelines typically involve matching spectra produced by the mass spectrometer to a theoretical spectra database, followed by matching the identified peptides back to parent-proteins. However, the nature of enzymatic digestion produces peptides that can be found in multiple proteins due to conservation or chance, presenting difficulties with protein and functional assignment.

Results: To combat this challenge, we developed pepFunk, a peptide-centric metaproteomic workflow focused on the analysis of human gut microbiome samples. Our workflow includes a curated peptide database annotated with Kyoto Encyclopedia of Genes and Genomes (KEGG) terms and a gene set variation analysis-inspired pathway enrichment adapted for peptide-level data. Analysis using our peptide-centric workflow is fast and highly correlated to a protein-centric analysis, and can identify more enriched KEGG pathways than analysis using protein-level data. Our workflow is open source and available as a web application or source code to be run locally.

Availability and implementation: pepFunk is available online as a web application at with open-source code available from


Supplementary information: Supplementary data are available at Bioinformatics online.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Computational Biology
  • Gastrointestinal Microbiome*
  • Humans
  • Peptides
  • Proteins
  • Software


  • Peptides
  • Proteins