title: Examination of segmental average mass spectra from liquid chromatography-tandem mass spectrometric (LC-MS/MS) data enables screening of multiple types of protein modifications.
abstract: It has been observed that a modified peptide and its non-modified counterpart, when analyzed with reverse phase liquid chromatography, usually share a very similar elution property [1-3]. Inasmuch as this property is common to many different types of protein modifications, we propose an informatics-based approach, featuring the generation of segmental average mass spectra ((sa)MS), that is capable of locating different types of modified peptides in two-dimensional liquid chromatography-mass spectrometric (LC-MS) data collected for regular protease digests from proteins in gels or solutions. To enable the localization of these peptides in the LC-MS map, we have implemented a set of computer programs, or the (sa)MS package, that perform the needed functions, including generating a complete set of segmental average mass spectra, compiling the peptide inventory from the Sequest/TurboSequest results, searching modified peptide candidates and annotating a tandem mass spectrum for final verification. Using ROCK2 as an example, our programs were applied to identify multiple types of modified peptides, such as phosphorylated and hexosylated ones, which particularly include those peptides that could have been ignored due to their peculiar fragmentation patterns and consequent low search scores. Hence, we demonstrate that, when complemented with peptide search algorithms, our approach and the entailed computer programs can add the sequence information needed for bolstering the confidence of data interpretation by the present analytical platforms and facilitate the mining of protein modification information out of complicated LC-MS/MS data.
authors.affiliation: Institute of Biochemistry & Molecular Biology, National Yang-Ming University, Taipei, 112, Taiwan.
authors.forename: Nai-Yu
authors.initials: NY
authors.lastname: Liu
authors.affiliation: Department of Life Science & Institute of Genome Science, National Yang-Ming University, Taipei, 112, Taiwan.
authors.forename: Hsiao-Hui
authors.initials: HH
authors.lastname: Lee
authors.affiliation: Institute of Biochemistry & Molecular Biology, National Yang-Ming University, Taipei, 112, Taiwan.
authors.forename: Zee-Fen
authors.initials: ZF
authors.lastname: Chang
authors.affiliation: Institute of Biochemistry & Molecular Biology, National Yang-Ming University, Taipei, 112, Taiwan; Proteomics Research Center, National Yang-Ming University, Taipei, 112, Taiwan. Electronic address: ygtsay@ym.edu.tw.
authors.forename: Yeou-Guang
authors.initials: YG
authors.lastname: Tsay
mesh_terms: D000595:Amino Acid Sequence; D000906:Antibodies / Q000276:immunology; D002851:Chromatography, High Pressure Liquid; D057809:HEK293 Cells; D006801:Humans; D047468:Immunoprecipitation; D008969:Molecular Sequence Data; D010748:Phosphopeptides / Q000032:analysis; D011499:Protein Processing, Post-Translational; D053719:Tandem Mass Spectrometry; D054460:rho-Associated Kinases / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism
chemical_list: D000906:Antibodies; D010748:Phosphopeptides; C515852:ROCK2 protein, human; D054460:rho-Associated Kinases
keywords: Bioinformatics; Liquid chromatography–tandem mass spectrometry (LC–MS/MS); Peptide elution; Protein modifications; Segmental average mass spectra