Fig. 2.

Residues 243–247 were affected by PTMs. A–E, the relative deuterium level is shown for five overlapping pepsin fragments covering heavy chain residues 242–254. The level of deuteration was monitored at 10 s and 1, 10, 60, and 240 min for each IgG1 variant. Solid black, native IgG1; solid gray, degalactosylated IgG1; dotted black, hypergalactosylated IgG1; solid red, deglycosylated IgG1; solid blue, methionine-oxidized IgG1; dotted blue, hypergalactosylated-methionine-oxidized IgG1. These traces are an average of four replicates with error bars at each time point ranging from ±0.05 to 0.15 Da. The sequence of residues 242–254 is shown at the bottom of the figure, and each different peptide (A–E) is depicted as a bar under the sequence. Note that because the values measured are relative deuterium levels the absolute value of deuterium at each time point cannot be compared with other measurements outside this data set (see “Experimental Procedures”). F, a zoomed-in view of the model structure of the IgG1 studied (Protein Data Bank codes 3FZU and 1HZH; see Ref. 22). The red spheres indicate the backbone nitrogen and amide hydrogen where the modifications produced differences in deuterium incorporation, and the green spheres represent Met-253 and Met-429. Glycans are colored cyan.