Refolding Record:
| Protein | |
|---|---|
| Protein Name | RNA polymerase |
| Abbreviated Name | RNAP |
| SCOP Family | RNA-polymerase beta-prime |
| Structure Notes | |
| Organism | Bordetella pertussis |
| UniProt Accession | P0A4E5 |
| SCOP Unique ID | n/a |
| Structure Solved | |
| Class | Multi-domain proteins (alpha and beta) |
| Molecularity | Monomer |
| Construct | |
|---|---|
| Full Length | n |
| Domain | Alpha Chain |
| Chimera | n/a |
| Variants | n/a |
| Chain Length | 328 |
| Molecular Weight | 36158.5 |
| Pi | 5.6 |
| Molecular Weight | 36158.5 |
| Disulphides | 0 |
| Full Sequence |
MSTQGFLKPRSIEVEPVGAHHAKIVMEPFERGYGHTLGNALRRILLSSMTGYAPTEVQMT
GVVHEYSTIAGVREDVVDILLNLKGVVFKLHNRDEVTLVLRKNGAGAVVASDIELPHDVE
IINPDHLICNLTDAGKIEMQVKVEKGRGYVPGNVRALSEDRTHTIGRIVLDASFSPVRRV
SYAVESARVEQRTDLDKLVLDIETNGVISPEEAVRQAARILMDQISVFAALEGAGDAYEP
PVRGTPQIDPVLLRPVDDLELTVRSANCLKAENIYYIGDLIQRTENELLKTPNLGRKSLN
EIKEVLAARGLTLGMKLENWPPLGLERP
|
| Notes | n/a |
| Expression | |
|---|---|
| Report | Boucher, P.E., Murakami, K., Ishihama, A., Stibitz, S (1997) J Bacteriol., 179, 1755-1763 |
| Project Aim | Protein-Protein Interaction Identification |
| Fusion | None |
| Protein Expression and Production | Protein expressed and purified in native conformation prior to denaturation and refolding. |
| Expression Host | None |
| Expression Strain | None |
| Expression Temp | 0.0 |
| Expression Time | 0 |
| Expression Vector | |
| Expression Protocol | |
| Method of Induction | Not Stated |
| Cell Density at Induction | OD = |
| Cell Disruption Method | None |
| Lytic Agent | None |
| Pre-Refolding Purification | Immunoaffinity Chromatography |
| Solubility | |
| Refolding | |
|---|---|
| Refolding Method | Dialysis |
| Wash Buffer | n/a |
| Solubilization Buffer | n/a |
| Refolding Buffer | 10mM TrisHCl, pH 7.9, 0.1mM EDTA, 0.2mM DTT, 100mM NaCl, 50% glycerol |
| Pre-Refolding Purification | Immunoaffinity Chromatography |
| Tag Cleaved | no tag |
| Refolding pH | 7.9 |
| Refolding Temperature | 4.0 |
| Protein Concentration | overnight |
| Refolding Time | n/a |
| Redox Agent | None |
| Redox Agent Concentration | n/a |
| Refolding Protocol | Protein was purified from B. pertussis and eluted from affinity column with 30% 2,3-butanediol and 0.75M ammonium sulfate in 50mM TrisHCl, pH 7.9, and 0.1mM EDTA. The eluted enzyme was dialyzed overnight against refolding buffer and aliquots containing 4.2 mg of protein were stored at -20degC. |
| Refolding Assay | Ligand Binding |
| Refolding Chaperones | None |
| Refolding Additives | Glycerol |
| Additives Concentration | 50% |
| Refolding Yield | n/a |
| Purity | n/a |
| Notes | n/a |