Table 1.
Steps of the isolation and purification of ribokinase, PRPP-synthetases, and APR-transferase
| Purification step | Volume, mL | Protein concentration, mg/mL | Total protein, mg |
|---|---|---|---|
| Ribokinase from Thermus sp. 2.9 | |||
| 1. Ultrasonic disintegration | 150* | 8.3 | 1245 |
| 2. Heat treatment | 136 | 1.7 | 231.2 |
| 3. Metal chelate chromatography | 30 | 1.9 | 57 |
| 4. Concentrating | 9 | 6 | 54 |
| 5. Gel filtration chromatography | 28.5 | 1.28 | 36.4 |
| 6. Concentrating | 2.6 | 11.7 | 30.4 |
| PRPP-synthetase 1 from T. thermophilus HB27 | |||
| 1. Ultrasonic disintegration | 150** | 10 | 1500 |
| 2. Affinity chromatography | 100 | 0.8 | 80 |
| 3. Concentrating | 13 | 5.7 | 74.1 |
| 4. Gel filtration | 50 | 1 | 50 |
| 5. Concentrating | 4 | 12.3 | 49.2 |
| PRPP-synthetase 2 from T. thermophilus HB27 | |||
| 1. Ultrasonic disintegration | 130** | 11.2 | 1456 |
| 2. Affinity chromatography | 125 | 0.6 | 75 |
| 3. Concentrating | 10.3 | 6.2 | 63.9 |
| 4. Gel filtration | 37.5 | 1.2 | 45 |
| 5. Concentrating | 3.4 | 12 | 40.8 |
| APR-transferase from T. thermophilus HB27 | |||
| 1. Ultrasonic disintegration | 157*** | 6 | 942 |
| 2. Heat treatment | 149 | 1.3 | 193.7 |
| 3. Gel filtration | 258 | 0.7 | 181 |
| 4. Anion exchange chromatography | 72 | 1.2 | 86.4 |
| 5. Hydrophobic interaction chromatography | 73 | 0.8 | 58.4 |
| 6. Concentrating | 11 | 5 | 55 |
| 7. Gel filtration chromatography | 52 | 0.87 | 45.2 |
| 8. Concentrating | 3.4 | 12.5 | 42.5 |
*From 5.8 L of culture.
**From 5 L of culture.
***From 6 L of culture.