| Â | Expected MW of Reduced Monomer | Expected MW of Oxidized Dimer | Purified Enzyme (Da) | Purifed Enzyme + 20 mM DTT (Da) |
|---|
Pf Trx-Px1 | 23702.08 | 47400.12 | 47456.56 (99%+) 23729.32 (trace) a | 23730.96 |
Pv Trx-Px1 | 23567.94 | 47131.84 | 47133.20 (major) 23568.94 (minor) | 23568.77 |
Py Trx-Px1 | 23534.86 | 47065.68 | 47066.49 (99%+) 23533.70 (trace) | 23533.70 |
Py Trx-Px1: Q7-L195 | 23234.40 | 46468.76 | 46470.60(99%+) 23234.49 (trace) | 23235.18 |
Pk Trx-Px1 | 24225.58 | 48447.12 | 48449.43 (99%+) 24226.10 (trace) | 24226.38 |
- The calculated MW in Da is determined from the amino acid sequence and includes the His6-tag incorporated into our constructs. The expected monomer MW is determined from the calculated monomer MW and subtracts the known E. coli post translational modification, namely clipping of the N-terminal Met (-131.19) of the His6-tag [26]. The expected MW of the oxidized dimer is calculated by doubling the expected MW weight of the monomer (clipping of the N-terminal Met accounted for) and subtracting 4 H (-4.04). aThe MW of the purified monomeric Pf Trx-Px-1 is off by 28 Da (and the dimer is off by twice this) which may be accounted for by one of the following: addition of ethyl addition, N, N-dimethylation of Arg or Lys, 2,4 bis-Trp-6,7-dione formation, or addition of formaldehyde (CHO)
