Table 1.
Biochemical properties of wild-type and mutant P4 proteins
| P4 | ϕ6 | ϕ8 | ϕ12 | ϕ13 | ϕ12 S292A |
ϕ12 Q278A |
ϕ12 Y288A |
ϕ12 ΔTTS202-204 |
ϕ12 TTS202-204LKK |
|---|---|---|---|---|---|---|---|---|---|
| Nucleotide binding | RNA binding L1 loop | ||||||||
| Mol weight (kDa) | 35 | 34.1 | 35.1 | 37.6 | 35.1 | 35.1 | 35.1 | 35.1 | 35.1 |
| Quaternary structure and stability | Hexamer7 (+ATP/ADP) | Hexamer (8) | Hexamer (26) | Hexamer (8) | Hexamer | Hexamer | Hexamer | Hexamer | Hexamer |
| Controlled ring opening (37) | Frequent ring opening (38) | ||||||||
| KM(ATP) mM | 0.19 ± 0.03 (8) | NA | 1.50 ± 0.04 (36) | 0.40 ± 0.05 (8) | 2.20 ± 0.5 | ||||
| kcat s-1 | 0.19 ± 0.06 (8) | 0.00 ± 0.05 (39) | 0.84 ± 0.12 (36) | 1.60 ± 0.05 (8) | 0 | 0.25 ± 0.13 | 0 | 0 | 0 |
| KM(ATP) mM with 1 mM polyC | ND | 0.17 ± 0.01 (39) | 0.49 ± 0.02 (36) | ND | 2.10 ± 0.1 | ||||
| kcat s-1 with 1 mM polyC | ND7 | 6.40 ± 0.20 (39) | 2.52 ± 0.07 (36) | ND | 0 | 0.96 ± 0.05 | 0 | 0 | 0 |
| NTP specificity | All (8) | All (8) | Purine base (26) | All (8) | |||||
| ssRNA binding | Kd > 1 Mm (8) | Kd<1 µM (8,39) | Kd> 1 mM (26,36) | Kd<1 µM (8) | |||||
| ssRNA translocation | Weak (7) | Strong | Weak (36) | Strong (8) | |||||
| Processive (36,39) | |||||||||
| COD (helicase) activity | Only in PC (8) | Strong (8) | None (8) | Weak (8) | |||||
| RNA binding site | ND | L1 (LKK) (37) | L2 (K241) (30) | ND | |||||