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protocol previously described [2]. The protein and monopotassium phosphoramidate wereused at the ratio 1:100 (w:w).All mass spectrometric experiments were performed on an Apex-Qe Ultra 7Tinstrument (Bruker Daltonics, Bremen, Germany) equipped with a dual ESI source. Allmass spectrometric experiments were performed on an Apex-Qe Ultra 7T and a heatedhollow cathode dispenser. The precursor ions were selected on the quadrupole and directedto the ICR cell where they were fragmented. The parameters were set as 150 ms for theECD pulse length and ECD bias was 0.8 V. Analysis of the obtained spectra was carriedout with the Biotools (Bruker) software. Obtained ECD spectra (Figure 1) allows theidentification of the numerous fragments of c n and (z+1) n . The number of identifiedfragments for diglycated and phosphorylated protein was 90 and 189 respectively whatindicate the high sequence coverage. The data analysis reveled that only one residue(His 68) undergoes phosphorylation. On the other hand the glycation reaction was lessspecific and all (or majority) lysine residues undergoes glycation.The ECD method was found useful for the localization of chemical modification sitesin glycated and phosphorylated ubiquitin. Its main advantage is a significant reduction ofthe neutral losses related to aminofructose (Fru) or phosphoramidate moiety (P).AcknowledgmentsSupported by Grant No. N N401 222734 from the MSHE (Poland).References1. Stefanowicz, P., Boratyński, J., Kanska, U., Petry, I., Szewczuk, Z. Acta Biochim. Pol. 48, 1137-1141 (2001).2. Kowalewska, K., Stefanowicz, P., Ruman, T., Fraczyk, T., Rode, W.K., Szewczuk, Z. BioscienceReports in press, doi:10.1042.3. Stefanowicz, P., Kijewska, M., Szewczuk, Z. J. Mass Spectrom. 44, 1047-1052 (2009).279