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Originally published In Press as doi:10.1074/mcp.T500034-MCP200 on January 25, 2006.
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Molecular & Cellular Proteomics 5:935-948, 2006.
© 2006 by The American Society for Biochemistry and Molecular Biology, Inc.

Technology

ModifiComb, a New Proteomic Tool for Mapping Substoichiometric Post-translational Modifications, Finding Novel Types of Modifications, and Fingerprinting Complex Protein Mixtures*

Mikhail M. Savitski{ddagger},§, Michael L. Nielsen{ddagger} and Roman A. Zubarev

From the Laboratory for Biological and Medical Mass Spectrometry, Uppsala University, S-75123 Uppsala, Sweden

A major challenge in proteomics is to fully identify and characterize the post-translational modification (PTM) patterns present at any given time in cells, tissues, and organisms. Here we present a fast and reliable method ("ModifiComb") for mapping hundreds types of PTMs at a time, including novel and unexpected PTMs. The high mass accuracy of Fourier transform mass spectrometry provides in many cases unique elemental composition of the PTM through the difference {Delta}M between the molecular masses of the modified and unmodified peptides, whereas the retention time difference {Delta}RT between their elution in reversed-phase liquid chromatography provides an additional dimension for PTM identification. Abundant sequence information obtained with complementary fragmentation techniques using ion-neutral collisions and electron capture often locates the modification to a single residue. The ({Delta}M, {Delta}RT) maps are representative of the proteome and its overall modification state and may be used for database-independent organism identification, comparative proteomic studies, and biomarker discovery. Examples of newly found modifications include +12.000 Da (+C atom) incorporation into proline residues of peptides from proline-rich proteins found in human saliva. This modification is hypothesized to increase the known activity of the peptide.

§ To whom correspondence should be addressed: Laboratory for Biological and Medical Mass Spectrometry, Uppsala University, Box 583, S-75123 Uppsala, Sweden. Tel.: 46-18-471-5729; Fax: 46-18-471-5729; E-mail: Mikhail.Savitski{at}bmms.uu.se


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