( b ...)Identification of Low-Level Oxidation Products in Calmodulin Using NanoSpray ,,, Ion Trap Mass Spectrometry

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Figure 2B. Full-scan MS³ spectrum of doubly-charged ion at m/z 1177.5 for peak at RT 45.87 min.

In contrast, the MS² spectrum of the doubly charged ion at m/z 1209.5 for the peak at RT = 49.45 min includes y and b-series ions, as well as the MetSO associated neutral loss for the b ions, indicating that the MetSO is located at the Nterminus of the peptide, e.g., Met₁₀₉ (Figure 3A and Figure 3A insert). MS³ analysis of the peak at m/z 1177.4 in the MS² spectrum substantiates oxidation of Met₁₀₉ through the detection of MetSO-specific neutral loss sequence ions of Nterminal origin (Figure 3B).

Similarly, MS² and MS³ experiments were conducted for the peak at RT = 56.31, and it was established that the corresponding peptide contained MetSO exclusively at Met₁₄₄. The extent and character of MetSO formation in CaM shown in this experiment is in agreement with results published previously².


Figure 3A. Full-scan MS² spectrum of doubly-charged ion at m/z 1209.5 for peak at RT 49.45 min.
Insert: Amino acid sequence of T₁₂ fragment indicating major C- and Nterminal fragment ions detected by full-scan MS².



Figure 3B. Full-scan MS³ spectrum of doubly-charged ion at m/z 1177.5 for peak at RT 49.45 min.

Conclusions

The identification of low-level oxidation products such as MetSO in proteins and peptides can be easily achieved using any Thermo Finnigan LCQ series ion trap mass spectrometer with MSⁿ capability. The combination of D
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