Table 2. MS analysis of DgkA.
| Sample | Attachment | Expected MW† (Da) | Observed MW‡ (Da) | Expected MW difference§ (Da) | Observed MW difference¶ (Da) |
|---|---|---|---|---|---|
| CM41 | None | 14187.48 | 14186.80 (80) | 0 | 0.68 |
| DM (1) | 14670.05 | 14669.58 (20) | 482.57 | 482.10 | |
| CM41DTNB | NTB2 | 14384.66 | 14384.45 | 197.18 | 196.97 |
| CM41EMP | None | 14187.48 | 14186.89 (39) | 0 | 0.59 |
| EMP (1) | 14416.13 | 14415.83 (31) | 228.65 | 228.35 | |
| EMP (2) | 14644.79 | 14644.45 (25) | 457.31 | 457.76 | |
| EMP (1), DM (1) | 14898.70 | 14898.84 (5) | 711.22 | 711.36 | |
| CM41CH3HgCl | None | 14187.48 | 14187.02 (43) | 0 | 0.46 |
| CH3HgCl (1) | 14402.11 | 14402.07 (39) | 214.63 | 214.59 | |
| CH3HgCl (2) | 14616.73 | 14616.93 (15) | 429.25 | 429.91 | |
| DM (1), CH3HgCl (1) | 14884.68 | 14884.41 (5) | 697.20 | 696.93 | |
| CM41-SeMet | SeMet | 14281.27 | 14281.61 (70) | 93.79 | 94.13 |
| SeMet, DM (1) | 14763.84 | 14764.19 (30) | 576.36 | 576.71 | |
| CM43 | None | 14159.42 | 14159.13 (76) | 0 | 0.29 |
| DM (1) | 14641.99 | 14642.16 (17) | 482.57 | 482.74 | |
| DM (2) | 15124.56 | 15124.43 (5) | 965.14 | 965.01 | |
| DM (3) | 15607.13 | 15607.91 (2) | 1447.71 | 1448.49 | |
| CM43DTNB | NTB2 (1) | 14356.60 | 14356.98 (90) | 197.18 | 197.56 |
| NTB2 (1), DM (1) | 14839.17 | 14839.20 (10) | 679.75 | 679.78 | |
| CM43EMP | None | 14159.42 | 14158.77 (35) | 0 | 0.65 |
| EMP (1) | 14388.07 | 14388.15 (23) | 228.65 | 228.73 | |
| EMP (2) | 14616.73 | 14616.54 (20) | 457.31 | 457.12 | |
| DM (1) | 14641.99 | 14642.72 (7) | 482.57 | 483.30 | |
| DM (1), EMP (1) | 14870.64 | 14870.78 (5) | 711.22 | 711.36 | |
| DM (1), EMP (2) | 15099.29 | 15099.19 (4) | 939.87 | 939.77 | |
| DM (1), EMP (1) | 15353.21 | 15352.85 (2) | 1193.79 | 1193.43 | |
| DM (2), EMP (2) | 15581.86 | 15580.00 (1) | 1422.44 | 1420.58 | |
| DM (3) | 15607.13 | 16606.40 (1) | 1447.23 | 1446.98 | |
| CM46 | None | 14187.48 | 14187.19 (70) | 0 | 0.29 |
| DM (1) | 14670.05 | 14669.70 (20) | 482.57 | 482.22 | |
| DM (2) | 15152.62 | 15152.00 (8) | 965.14 | 964.52 | |
| DM (3) | 15635.19 | 15635.00 (2) | 1447.71 | 1447.52 | |
| CM46DTNB | NTB2 (1) | 14384.66 | 14384.88 (80) | 197.18 | 197.40 |
| NTB2 (1), DM (1) | 14867.23 | 14867.06 (16) | 679.75 | 679.58 | |
| NTB2 (1), DM (2) | 15349.80 | 15349.30 (4) | 1162.32 | 1161.82 | |
| CM46EMP | None | 14187.48 | 14187.71 (49) | 0 | 0.23 |
| EMP (1) | 14670.05 | 14416.65 (20) | 228.65 | 229.17 | |
| EMP (2) | 14644.79 | 14645.15 (17) | 457.31 | 457.67 | |
| DM (1) | 14670.05 | 14671.15 (9) | 482.57 | 483.67 | |
| DM (1), EMP (1) | 14898.70 | 14899.78 (2) | 711.22 | 712.30 | |
| DM (1), EMP (2) | 15127.35 | 15127.60 (2) | 939.87 | 940.12 | |
| DM (2) | 15152.62 | 15153.59 (1) | 965.14 | 966.11 |
The expected MW of nonlabelled protein was calculated based on the amino-acid composition of the DgkA construct with the N-terminal Met missing as a result of processing in vivo (Frottin et al., 2006 ▶; Hopper et al., 2013 ▶). The expected MWs for mercury/Ellman’s reagent modifications were calculated based on a reaction in which the added chemical groups replace the hydrogen of the thiol group. For SeMet-labelled protein, the value was calculated by replacing the S atom in the two methionine residues with selenium.
The values in parentheses are the percentages of each species, which were calculated based on peak height, assuming that the samples with and without labelling behave in the same manner during the MS experiment.
Refers to the MW difference between the labelled and unlabelled protein.
The MW difference was calculated using the observed MW subtracted from the theoretical MW of the unlabelled protein.