Mass fingerprinting of complex mixtures: protein inference from high-resolution peptide masses and predicted retention times.
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.
Science for Life Laboratory, Department of Biochemistry and Biophysics, Stockholm University , Tomtebodavägen 23A, 17165 Solna, Sweden.
Luminita
L
Moruz
Michael R
MR
Hoopmann
Magnus
M
Rosenlund
Viktor
V
Granholm
Robert L
RL
Moritz
Lukas
L
Käll
D000595:Amino Acid Sequence; D002853:Chromatography, Liquid; D045424:Complex Mixtures / Q000737:chemistry*; D006801:Humans; D015233:Models, Statistical; D058977:Molecular Sequence Annotation; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification*; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification*; D059748:Proteolysis; D012441:Saccharomyces cerevisiae / Q000737:chemistry; D013194:Staining and Labeling / Q000379:methods; D053719:Tandem Mass Spectrometry / Q000706:statistics & numerical data*
D045424:Complex Mixtures; D010446:Peptide Fragments; D011506:Proteins