Mass fingerprinting of complex mixtures: protein inference from high-resolution peptide masses and predicted retention times

J Proteome Res. 2013 Dec 6;12(12):5730-41. doi: 10.1021/pr400705q. Epub 2013 Oct 11.

Abstract

In typical shotgun experiments, the mass spectrometer records the masses of a large set of ionized analytes but fragments only a fraction of them. In the subsequent analyses, normally only the fragmented ions are used to compile a set of peptide identifications, while the unfragmented ones are disregarded. In this work, we show how the unfragmented ions, here denoted MS1-features, can be used to increase the confidence of the proteins identified in shotgun experiments. Specifically, we propose the usage of in silico mass tags, where the observed MS1-features are matched against de novo predicted masses and retention times for all peptides derived from a sequence database. We present a statistical model to assign protein-level probabilities based on the MS1-features and combine this data with the fragmentation spectra. Our approach was evaluated for two triplicate data sets from yeast and human, respectively, leading to up to 7% more protein identifications at a fixed protein-level false discovery rate of 1%. The additional protein identifications were validated both in the context of the mass spectrometry data and by examining their estimated transcript levels generated using RNA-Seq. The proposed method is reproducible, straightforward to apply, and can even be used to reanalyze and increase the yield of existing data sets.

Publication types

  • Research Support, American Recovery and Reinvestment Act
  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Chromatography, Liquid
  • Complex Mixtures / chemistry*
  • Humans
  • Models, Statistical*
  • Molecular Sequence Annotation
  • Molecular Sequence Data
  • Peptide Fragments / chemistry
  • Peptide Fragments / isolation & purification*
  • Proteins / chemistry
  • Proteins / isolation & purification*
  • Proteolysis
  • Saccharomyces cerevisiae / chemistry
  • Staining and Labeling / methods
  • Tandem Mass Spectrometry / statistics & numerical data*

Substances

  • Complex Mixtures
  • Peptide Fragments
  • Proteins