title: Characterization of protein serotonylation via bioorthogonal labeling and enrichment.
abstract: Protein serotonylation is a transglutaminase-mediated phenomenon whose biological mechanism of protein serotonylation is not yet fully understood, as the complete profiling of serotonylation targets in a proteome remains a critical challenge to date. Utilizing an alkyne-functionalized serotonin derivative bioorthogonally coupled to a cleavable linker, we developed a method to selectively enrich serotonylated proteins in a complex sample. With online nanoflow liquid chromatography and LTQ-Orbitrap Velos hybrid mass spectrometer detection, we identified 46 proteins with 50 serotonylation sites at their glutamine residues. Mass spectrometric analysis also generated direct residue-level evidence of various biological processes such as transglutaminase-chaperon interactions as well as actin assembly. An enrichment workflow utilizing click chemistry and on-bead digestion allowed us to achieve site-specific identification of protein serotonylation by mass spectrometry, and results obtained hereby also provided a great foundation in the elucidation of the true roles of protein serotonylation in biological systems.
authors: Institute of Biological Chemistry, ‡Core Facilities for Protein Structural Analysis, and §Chemical Biology and Molecular Biophysics, Taiwan International Graduate Program, Academia Sinica , 128 Academia Road Section 2, Taipei 11529, Taiwan.
authors: Jason Ching-Yao
authors: JC
authors: Lin
authors: Chi-Chi
authors: CC
authors: Chou
authors: Zhijay
authors: Z
authors: Tu
authors: Lun-Fu
authors: LF
authors: Yeh
authors: Shang-Chuen
authors: SC
authors: Wu
authors: Kay-Hooi
authors: KH
authors: Khoo
authors: Chun-Hung
authors: CH
authors: Lin
mesh_terms: D002853:Chromatography, Liquid / Q000379:methods; D057930:Click Chemistry; D005973:Glutamine / Q000378:metabolism; D013058:Mass Spectrometry / Q000379:methods; D011506:Proteins / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*; D012701:Serotonin / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D013194:Staining and Labeling
chemical_list: D011506:Proteins; D005973:Glutamine; D012701:Serotonin