TABLE 1.
Purification of recombinant Streptomyces sp. strain C5 from S. lividans TK24(pANT195)
| Fraction | Total protein (mg) | Specific content of P-450 (nmol/mg) | C-13 hydroxylation activity (pmol/min/mg) | Ratio of 13-hydroxylation activity per nmol of heme | Recovery (%)a | Purification (fold)b |
|---|---|---|---|---|---|---|
| Crude extract | 4,670 | 0.07 | 4.4 | 61 | 100 | 1 |
| Ammonium sulfate | 2,850 | 0.09 | 8.6 | 94 | 77 | 2.0 |
| Q-Sepharose Fast Flow | 906 | 0.23 | 19.3 | 84 | 62 | 4.5 |
| Econo-Pac High S | 720 | 0.29 | 26.1 | 90 | 63 | 6.1 |
| Phenyl Sepharose CL-4B | 60 | 1.33 | 490 | 368 | 24 | 114 |
| Superose 6 (FPLC) | 7.2 | 1.28 | 603 | 471 | 2.8 | 140 |
a
Based on P-450 heme content (i.e., active enzyme).
b
Based on the specific activity of 13-oxidation of 13-dihydrodaunorubicin to daunorubicin.