Cloned (Comment) | Organism |
---|---|
recombinant enzyme P2 expression in Escherichia coli strain BL21 (DE3) | Pseudomonas phage phi12 |
Crystallization (Comment) | Organism |
---|---|
enzyme in two crystal forms A and B, method screening, 300 nl of 6-20 mg/ml protein in 50 mM Tris, 100 mM NaCl, and 2 mM DTT, pH 8.0, are mixed with reservoir solution containing 4.0 M sodium formate, 0.2 M magnesium formate, and 20% w/v PEG 3350, in hanging drops at 20°C, or with 4.3 M NaCl, 0.1 M HEPES, and 30 mM tri-glycine in sitting drops at room temperature, X-ray diffraction structure determination and analysis at resolutions of 1.7 A and 2.1 A, respectively. Form A contains Mg2+ bound at a site that deviates from the canonical non-catalytic position seen in form B | Pseudomonas phage phi12 |
Protein Variants | Comment | Organism |
---|---|---|
additional information | temperature sensitivity of a ts-mutant | Pseudomonas phage phi12 |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
Ca2+ | inhibits catalyis | Pseudomonas phage phi12 |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Mg2+ | required for catalyis | Pseudomonas phage phi12 | |
additional information | Mn2+ is not required for catalyis | Pseudomonas phage phi12 |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
nucleoside triphosphate + RNAn | Pseudomonas phage phi12 | - |
diphosphate + RNAn+1 | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Pseudomonas phage phi12 | Q94M06 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant enzyme P2 from Escherichia coli strain BL21 (DE3) by anion exchange chromatography, gel filtration, dialysis, and ultrafiltration | Pseudomonas phage phi12 |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
additional information | template ssRNA accesses the active site through a tunnel. The phi12 P2 conformation does not favor RNA binding | Pseudomonas phage phi12 | ? | - |
? | |
nucleoside triphosphate + RNAn | - |
Pseudomonas phage phi12 | diphosphate + RNAn+1 | - |
? |
Synonyms | Comment | Organism |
---|---|---|
RDRP | - |
Pseudomonas phage phi12 |
self-priming RdRp | - |
Pseudomonas phage phi12 |
General Information | Comment | Organism |
---|---|---|
additional information | structure of P2, the self-priming RdRp from cystovirus phi12, crystal structure analysis, overview. The tunnel through which template ssRNA accesses the active site is partially occluded by a flexible loop; this feature, along with sub-optimal positioning of other structural elements that prevent the formation of a stable initiation complex, indicate an inactive conformation in crystallo | Pseudomonas phage phi12 |
physiological function | P2, an RNA-directed polymerase (RdRp), is the critical component of a four-protein polymerase complex (PX) that includes in addition to P2, the major capsid protein, P1, a packaging NTPase, P4, and an accessory protein, P71. P2 performs the dual tasks of transcription and replication de novo without the use of a primer (i. e. P2 is a self-priming RdRp), and plays a critical role in the cystoviral life cycle | Pseudomonas phage phi12 |