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Proteomics

Proteomics

A proteome is the entire set of proteins that a cell type produces. We can study proteomes using the knowledge of genomes because genes code for mRNAs, and the mRNAs encode proteins. Although mRNA analysis is a step in the right direction, not all mRNAs are translated into proteins.
Proteomics is the study of proteomes' function. It involves the large-scale systematic study of the proteome to denote the protein complement expressed by a genome. Scientist Mark Wilkins coined the term...

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Related Experiment Video

Updated: Feb 18, 2026

Phosphopeptide Enrichment Coupled with Label-free Quantitative Mass Spectrometry to Investigate the Phosphoproteome in Prostate Cancer

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Quantifying Missing (Phospho)Proteome Regions with the Broad-Specificity Protease Subtilisin.

Humberto Gonczarowska-Jorge^1,2, Stefan Loroch¹, Margherita Dell'Aica¹

¹Leibniz-Institut für Analytische Wissenschaften - ISAS - e.V. , 44227 Dortmund, Germany.

Analytical Chemistry

|November 15, 2017

Summary

Subtilisin, a broad-specificity protease, significantly enhances human proteome mapping by accessing previously hidden regions, especially proline-rich areas. It also aids in profiling post-translational modifications and offers fast, reproducible protein quantification.

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Quantitative Phosphoproteomics in Fatty Acid Stimulated Saccharomyces cerevisiae

Quantitative Phosphoproteomics in Fatty Acid Stimulated Saccharomyces cerevisiae

Published on: October 12, 2009

Area of Science:

Proteomics
Mass Spectrometry
Enzymology

Background:

Human proteome mapping efforts using mass spectrometry achieve <50% sequence coverage.
Protease-resistant domains, nonunique peptides, and ionization issues limit proteome coverage.
Complementary proteases like Glu-C and Asp-N improve coverage but have limitations.

Purpose of the Study:

To evaluate the broad-specificity protease subtilisin for human proteome mapping.
To assess subtilisin's digestion efficiency, reproducibility, and comparison to trypsin.
To explore subtilisin's utility in identifying novel proteomic regions and post-translational modifications.

Main Methods:

Systematic evaluation of subtilisin digestion efficiency and reproducibility on cell lysates.
Comparison of subtilisin and trypsin digestion for protein identification and sequence coverage.
Analysis of post-translational modification sites, particularly phosphorylation, identified by each enzyme.
In silico digestion analysis to assess accessibility of modification sites.

Main Results:

Subtilisin enables reproducible near-complete cell lysate digestion in 1-5 minutes.
Subtilisin identified 6807 proteins, while trypsin identified 8363 (1% protein FDR).
Subtilisin significantly improved coverage of missing and proline-rich proteome areas.
37% of 14,628 phosphorylation sites were exclusive to subtilisin, with 926 inaccessible by other enzymes.
Subtilisin demonstrated comparable precision to trypsin for in-depth quantification.

Conclusions:

Subtilisin is a valuable tool for expanding proteome coverage, particularly for challenging regions.
It is highly effective for system-wide profiling of post-translational modification sites.
Subtilisin offers a rapid and reproducible method for proteomic analysis and quantification.