Table 1. Cleavage of the histidine tag from OBP44a by TEV protease.
(A) Effect of TCEP in the digestion buffer on the integrity of disulfide bonds. (B) Cleavage efficiency under different ratios of TEV to OBP44a, without reducing agent in the buffer at 4 °C overnight.
| A | ||||||
|---|---|---|---|---|---|---|
| Tube # | TCEP (mM) | Tev: OBP44a | Temperature and length | Mass measured compared to calculated mass with disulfide bonds | ||
| 1 | 0 | 1:10 | 24 °C 3 hours | 0.09 | ||
| 2 | 0 | 1:100 | 24 °C 3 hours | 0.17 | ||
| 3 | 0.1 | 1:10 | 24 °C 3 hours | 1.27 | ||
| 4 | 0.1 | 1:100 | 24 °C 3 hours | 1.22 | ||
| 5 | 0 | 1:10 | 4 °C overnight | 0.14 | ||
| 6 | 0 | 1:100 | 4 °C overnight | 0.14 | ||
| 7 | 0.1 | 1:10 | 4 °C overnight | 1.26 | ||
| 8 | 0.1 | 1:100 | 4 °C overnight | 1.21 | ||
| B | |
|---|---|
| Tev:OBP44a | Cleavage % |
| 1:100 | 60% |
| 1:50 | 77% |
| 1:25 | 95% |
| 1:10 | 100% |