Improving SRM assay development: a global comparison between triple quadrupole, ion trap, and higher energy CID peptide fragmentation spectra.

In proteomics, selected reaction monitoring (SRM) is rapidly gaining importance for targeted protein quantification. The triple quadrupole mass analyzers used in SRM assays allow for levels of specificity and sensitivity hard to accomplish by more standard shotgun proteomics experiments. Often, an SRM assay is built by in silico prediction of transitions and/or extraction of peptide precursor and fragment ions from a spectral library. Spectral libraries are typically generated from nonideal ion trap based shotgun proteomics experiments or synthetic peptide libraries, consuming considerable time and effort. Here, we investigate the usability of beam type CID (or "higher energy CID" (HCD)) peptide fragmentation spectra, as acquired using an Orbitrap Velos, to facilitate SRM assay development. Therefore, peptide fragmentation spectra, obtained by ion-trap CID, triple-quadrupole CID (QqQ-CID) and Orbitrap HCD, originating from digested cellular lysates, were compared. Spectral comparison and a dedicated correlation algorithm indicated significantly higher similarity between QqQ-CID and HCD fragmentation spectra than between QqQ-CID and ion trap-CID spectra. SRM transitions generated using a constructed HCD spectral library increased SRM assay sensitivity up to 2-fold, when compared to the use of a library created from more conventionally used ion trap-CID spectra, showing that HCD spectra can assist SRM assay development.

[1]  Craig D Wenger,et al.  Analysis of tandem mass spectra by FTMS for improved large-scale proteomics with superior protein quantification. , 2010, Analytical chemistry.

[2]  Rong Wang,et al.  The need for a public proteomics repository , 2004, Nature Biotechnology.

[3]  S. Ficarro,et al.  Optimized orbitrap HCD for quantitative analysis of phosphopeptides , 2009, Journal of the American Society for Mass Spectrometry.

[4]  Nichole L. King,et al.  The PeptideAtlas Project , 2010, Proteome Bioinformatics.

[5]  P. Thibault,et al.  Characterization of a high-pressure quadrupole collision cell for low-energy collision-indneed dissociation , 1994, Journal of the American Society for Mass Spectrometry.

[6]  M. Mann,et al.  Higher-energy C-trap dissociation for peptide modification analysis , 2007, Nature Methods.

[7]  Gary M. Hieftje,et al.  Computer Identification of Infrared Spectra by Correlation-Based File Searching, , 1978 .

[8]  Susan E Abbatiello,et al.  Effect of collision energy optimization on the measurement of peptides by selected reaction monitoring (SRM) mass spectrometry. , 2010, Analytical chemistry.

[9]  Matthias Mann,et al.  A Dual Pressure Linear Ion Trap Orbitrap Instrument with Very High Sequencing Speed* , 2009, Molecular & Cellular Proteomics.

[10]  Steven P Gygi,et al.  The absolute quantification strategy: a general procedure for the quantification of proteins and post-translational modifications. , 2005, Methods.

[11]  D. Lauffenburger,et al.  Multiple reaction monitoring for robust quantitative proteomic analysis of cellular signaling networks , 2007, Proceedings of the National Academy of Sciences.

[12]  Brendan MacLean,et al.  Bioinformatics Applications Note Gene Expression Skyline: an Open Source Document Editor for Creating and Analyzing Targeted Proteomics Experiments , 2022 .

[13]  Joost W Gouw,et al.  Highly robust, automated, and sensitive online TiO2-based phosphoproteomics applied to study endogenous phosphorylation in Drosophila melanogaster. , 2008, Journal of proteome research.

[14]  Gennifer E. Merrihew,et al.  Expediting the development of targeted SRM assays: using data from shotgun proteomics to automate method development. , 2009, Journal of proteome research.

[15]  Ruedi Aebersold,et al.  High-throughput generation of selected reaction-monitoring assays for proteins and proteomes , 2010, Nature Methods.

[16]  J. Yates,et al.  An approach to correlate tandem mass spectral data of peptides with amino acid sequences in a protein database , 1994, Journal of the American Society for Mass Spectrometry.

[17]  S. Mohammed,et al.  Improved peptide identification by targeted fragmentation using CID, HCD and ETD on an LTQ-Orbitrap Velos. , 2011, Journal of proteome research.

[18]  William Stafford Noble,et al.  Analysis of peptide MS/MS spectra from large-scale proteomics experiments using spectrum libraries. , 2006, Analytical chemistry.

[19]  Lukas N. Mueller,et al.  Full Dynamic Range Proteome Analysis of S. cerevisiae by Targeted Proteomics , 2009, Cell.

[20]  S. Hubbard,et al.  Observations on the detection of b- and y-type ions in the collisionally activated decomposition spectra of protonated peptides. , 2009, Rapid communications in mass spectrometry : RCM.

[21]  Robertson Craig,et al.  The use of proteotypic peptide libraries for protein identification. , 2005, Rapid communications in mass spectrometry : RCM.

[22]  S. A. McLuckey,et al.  Ion trap versus low-energy beam-type collision-induced dissociation of protonated ubiquitin ions. , 2006, Analytical chemistry.

[23]  Olga Vitek,et al.  Correlation between y-type ions observed in ion trap and triple quadrupole mass spectrometers. , 2009, Journal of proteome research.

[24]  S. Mohammed,et al.  Strong cation exchange (SCX) based analytical methods for the targeted analysis of protein post-translational modifications. , 2011, Current opinion in biotechnology.

[25]  Robertson Craig,et al.  Open source system for analyzing, validating, and storing protein identification data. , 2004, Journal of proteome research.

[26]  Jeroen Krijgsveld,et al.  Lys-N and trypsin cover complementary parts of the phosphoproteome in a refined SCX-based approach. , 2009, Analytical chemistry.

[27]  S. Gygi,et al.  Absolute quantification of proteins and phosphoproteins from cell lysates by tandem MS , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[28]  J. Yates,et al.  Method to compare collision-induced dissociation spectra of peptides: potential for library searching and subtractive analysis. , 1998, Analytical chemistry.

[29]  M. Mann,et al.  Feasibility of large-scale phosphoproteomics with higher energy collisional dissociation fragmentation. , 2010, Journal of proteome research.

[30]  Sándor Suhai,et al.  Fragmentation Pathways of Protonated Peptides , 2006 .