Vibrational Signatures of Protonated, Phosphorylated Amino Acids in the Gas Phase

Structural characterization of protonated phosphorylated serine, threonine, and tyrosine was performed using mid-infrared multiple photon dissociation (IRMPD) spectroscopy and density functional theory (DFT) calculations. The ions were generated and analyzed by an external electrospray source coupled to a Paul ion-trap type mass spectrometer. Their fragmentation was induced by the resonant absorption of multiple photons from a tunable free electron laser (FEL) beam. IRMPD spectra were recorded in the 900−1850 cm-1 energy range and compared to the corresponding computed IR spectra. On the basis of the frequency and intensity of two independent bands in the 900−1400 cm-1 energy range, it is possible to identify the phosphorylated residue. IRMPD spectra for a 12-residue fragment of stathmin in its phosphorylated and nonphosphorylated forms were also recorded in the 800−1400 cm-1 energy range. The lack of spectral congestion in the 900−1300 cm-1 region makes their distinction facile. Our results show that IRMPD spectroscopy may became a valuable tool for structural characterization of small phosphorylated peptides.