title: Opening a SWATH Window on Posttranslational Modifications: Automated Pursuit of Modified Peptides.
abstract: Posttranslational modifications of proteins play an important role in biology. For example, phosphorylation is a key component in signal transduction in all three domains of life, and histones can be modified in such a variety of ways that a histone code for gene regulation has been proposed. Shotgun proteomics is commonly used to identify posttranslational modifications as well as chemical modifications from sample processing. However, it favors the detection of abundant peptides over the repertoire presented, and the data analysis usually requires advance specification of modification masses and target amino acids, their number constrained by available computational resources. Recent advances in data independent acquisition mass spectrometry technologies such as SWATH-MS enable a deeper recording of the peptide contents of samples, including peptides with modifications. Here, we present a novel approach that applies the power of SWATH-MS analysis to the automated pursuit of modified peptides. With the new SWATHProphet(PTM) functionality added to the open source SWATHProphet software, precursor ions consistent with a modification are identified along with the mass and localization of the modification in the peptide sequence in a sensitive and unrestricted manner without the need to anticipate the modifications in advance. Using this method, we demonstrate the detection of a wide assortment of modified peptides, many unanticipated, in samples containing unpurified synthetic peptides and human urine, as well as in phospho-enriched human tissue culture cell samples.
authors.affiliation: From the §Institute for Systems Biology, 401 Terry Avenue North, Seattle, WA 98109.
authors.forename: Andrew
authors.initials: A
authors.lastname: Keller
authors.affiliation: From the §Institute for Systems Biology, 401 Terry Avenue North, Seattle, WA 98109.
authors.forename: Samuel L
authors.initials: SL
authors.lastname: Bader
authors.affiliation: From the §Institute for Systems Biology, 401 Terry Avenue North, Seattle, WA 98109.
authors.forename: Ulrike
authors.initials: U
authors.lastname: Kusebauch
authors.affiliation: From the §Institute for Systems Biology, 401 Terry Avenue North, Seattle, WA 98109.
authors.forename: David
authors.initials: D
authors.lastname: Shteynberg
authors.affiliation: From the §Institute for Systems Biology, 401 Terry Avenue North, Seattle, WA 98109.
authors.forename: Leroy
authors.initials: L
authors.lastname: Hood
authors.affiliation: From the §Institute for Systems Biology, 401 Terry Avenue North, Seattle, WA 98109 robert.moritz@systemsbiology.org.
authors.forename: Robert L
authors.initials: RL
authors.lastname: Moritz
mesh_terms: D045744:Cell Line, Tumor; D019295:Computational Biology / Q000379:methods; D006657:Histones / Q000737:chemistry; D006801:Humans; D010455:Peptides / Q000737:chemistry* / Q000652:urine; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D011506:Proteins / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D012984:Software; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D006657:Histones; D010455:Peptides; D011506:Proteins