TABLE 1.
Summary of the purification processes for the recombinant C. virescens (rCviUPO) and D. caldariorum (rDcaUPO) UPOs from E. coli cultures, using ABTS or veratryl alcohol as the substratea
| UPO and fraction | Vol (ml) | Protein (mg) | Activity (total U) | Sp act (U · mg−1) | Yield (%) | Purification (fold) |
|---|---|---|---|---|---|---|
| rCviUPOb | ||||||
| Cell lysate | 60 | 6,650 | 1,150 (60) | 0.17 (0.01) | 100 | 1 |
| Cation exchange | 30 | 59 | 292 (36) | 4.91 (0.61) | 25 (60) | 28 (67) |
| Ultrafiltrate | 1 | 56 | 240 (39) | 4.29 (0.69) | 21 (64) | 25 (76) |
| Superdex 75 | 8 | 8 | 309 (14) | 38.18 (1.70) | 27 (23) | 221 (189) |
| rDcaUPOc | ||||||
| Cell lysate | 100 | 4,230 | 243 (54) | 0.06 (0.01) | 100 | 1 |
| Metal affinity | 70 | 29 | 131 (41) | 4.48 (1.39) | 54 (75) | 78 (109) |
| Ultrafiltrate | 1 | 28 | 167 (36) | 5.90 (1.29) | 69 (68) | 103 (101) |
| Superdex 75 | 10 | 4 | 31 (7) | 7.68 (1.62) | 13 (13) | 125 (128) |
a
Activity, yield, and purification were measured with 1 mM ABTS in 0.1 M tartrate (pH 3) in the presence of 1 mM H2O2 or with 5 mM veratryl alcohol in 0.1 M Tris (pH 7.4); values for veratryl alcohol are shown in parentheses.
b
From an 8-liter E. coli culture.
c
From a 10-liter E. coli culture.