FIGURE 8.

Marfey's reagent derivatization of the VY* dipeptide isolated from reactions containing natural abundance MftA, [¹³C₉,¹⁵N]Tyr³⁰ MftA, or [¹³C₁₄,¹⁵N₂]²⁹VY³⁰ MftA. A, a new species (●) is observed in the extracted ion chromatograms that elutes at approximately 13 min when monitoring for the Marfey's reagent-derivatized VY* dipeptide with either no (unlabeled) (m/z = 486.8–487.7), ¹³C₈,¹⁵N (m/z = 495.8–496.8), or ¹³C₁₃,¹⁵N₂ (m/z = 501.8–502.8) isotopic enrichment. The peak at 13 min is only observed in reactions containing MftE and SAM (+SAM +MftE) and is not observed when SAM is removed (−SAM +MftE). B, full mass spectra of the species that elutes at approximately 13 min in the extracted ion chromatograms showing isotope-sensitive peaks for the [M + H]⁺ at m/z 487.1928, 496.2165, and 502.2297 with unlabeled MftA, [¹³C₈,¹⁵N]Tyr³⁰ MftA, or [¹³C₁₃,¹⁵N₂]²⁹VY³⁰ MftA. C, HCD fragmentation mass spectra of the [M + H]⁺ parent ion peak in the mass spectra shown in B. Possible structures for the fragments are shown in supplemental Fig. S5. D and E, the two possible structures and theoretical masses of the Marfey's reagent-derivatized VY* dipeptide based on the HCD fragmentation results with the natural abundance and isotopically enriched MftA peptides.