TABLE 2.
Average molecular mass determinations of 2-KPCC using LC-MS and two spectral deconvolution algorithms
Under the conditions utilized for analysis, 2-KPCC was observed as the FAD-depleted monomer.
| Sample | Theoretical average molecular mass | Measured average molecular mass by deconvolution algorithm | Mass accuracy (Meas-Theor) (for BIOMASS 3.1) | Mass change with BES modification (2-KPCC + BES + DTT) − (2-KPCC + BES) (for BIOMASS 3.1) | ||
|---|---|---|---|---|---|---|
| BIOMASS 3.1 | ProMass 2.0 | |||||
| M | M ± S.D. | ΔDa | Dppm | ΔDa | ||
| Oxidized 2-KPCC (run 1) | 57,345.6a | 57,365.0 | 57,362.1 ± 12.9 | +19.4 | 338 | |
| Oxidized 2-KPCC (run 2) | 57,345.6a | 57,358.0 | 57,355.7 ± 6.9 | +12.4 | 216 | |
| Oxidized 2-KPCC + BES | 57,345.6b | 57,361.0 | 57,358.4 ± 11.0 | +15.4 | 268 | |
| DTT-reduced 2-KPCC + BES | 57,454.7c | 57,470.0 | 57,463.3 ± 10.2 | +15.3 | 266 | 109.0 |
a Calculated using the ExPASy Compute pI/MW tool using the 2-KPCC protein sequence (Q56839) minus the average atomic mass of two hydrogens.
b Calculated awith no BES modification assumed.
c The average mass with BES modification of a single cysteine (+C2H4SO3 = 108.1; calculated using the average atomic masses of the elements, with the sum being rounded to the nearest 0.1 Da). The calculation assumed the SO3 moiety of BES and the Cys87 thiol of 2-KPCC to be deprotonated and protonated, respectively, under the conditions used for the analysis (in a solvent containing 0.1% (v/v) formic acid).