Refolding Record:
| Protein | |
|---|---|
| Protein Name | Quinolinate synthetase |
| Abbreviated Name | QS |
| SCOP Family | Unknown |
| Structure Notes | |
| Organism | Escherichia coli |
| UniProt Accession | P11458 |
| SCOP Unique ID | n/a |
| Structure Solved | |
| Class | Unknown |
| Molecularity | Monomer |
| Construct | |
|---|---|
| Full Length | y |
| Domain | n/a |
| Chimera | n/a |
| Variants | n/a |
| Chain Length | 352 |
| Molecular Weight | 38240.8 |
| Pi | 5.19323 |
| Molecular Weight | 38240.8 |
| Disulphides | Unknown |
| Full Sequence |
MSVMFDPDTAIYPFPPKPTPLSIDEKAYYREKIKRLLKERNAVMVAHYYTDPEIQQLAEE
TGGCISDSLEMARFGAKHPASTLLVAGVRFMGETAKILSPEKTILMPTLQAECSLDLGCP
VEEFNAFCDAHPDRTVVVYANTSAAVKARADWVVTSSIAVELIDHLDSLGEKIIWAPDKH
LGRYVQKQTGGDILCWQGACIVHDEFKTQALTRLQEEYPDAAILVHPESPQAIVDMADAV
GSTSQLIAAAKTLPHQRLIVATDRGIFYKMQQAVPDKELLEAPTAGEGATCRSCAHCPWM
AMNGLQAIAEALEQEGSNHEVHVDERLRERALVPLNRMLDFAATLRG
|
| Notes | n/a |
| Expression | |
|---|---|
| Report | Ceciliani F, Caramori T, Ronchi S, Tedeschi G, Mortarino M, Galizzi A. (2000) Protein Expression and Purification, 18, 64-70 |
| Project Aim | Functional Studies |
| Fusion | None |
| Protein Expression and Production | Protein recombinantly expressed as and refolded from inclusion bodies. |
| Expression Host | Escherichia coli |
| Expression Strain | TOP10 |
| Expression Temp | 37.0 |
| Expression Time | 5h |
| Expression Vector | pTrcHisA |
| Expression Protocol | unknown |
| Method of Induction | Not Stated |
| Cell Density at Induction | OD = |
| Cell Disruption Method | Sonication |
| Lytic Agent | None |
| Pre-Refolding Purification | None |
| Solubility | insoluble |
| Refolding | |
|---|---|
| Refolding Method | Dialysis |
| Wash Buffer | 50mM Tris-HCl pH 8.0 |
| Solubilization Buffer | 50mM Tris-HCl, 10mM EDTA, 0.1mM PMSF, 4M urea, pH 8.0 |
| Refolding Buffer | 50mM Tris-HCl, 10mM EDTA, 10% glycerol |
| Pre-Refolding Purification | None |
| Tag Cleaved | no tag |
| Refolding pH | 8.0 |
| Refolding Temperature | 4.0 |
| Protein Concentration | 1mg/ml |
| Refolding Time | 24h |
| Redox Agent | None |
| Redox Agent Concentration | n/a |
| Refolding Protocol | The solubilized material was dislysed against 50mM Tris-HCl, 10mM EDTA, 10% glycerol and 2M urea pH 8.0 for 5h. Subsequently the refolding buffer was changed to 50mM Tris-HCl, 10mM EDTA, 10% glycerol pH 8.0. After 24h the solution was centrifuged and the supernatant was further purified by anio exchange chromotography. |
| Refolding Assay | Enzyme activity |
| Refolding Chaperones | None |
| Refolding Additives | None |
| Additives Concentration | NULL |
| Refolding Yield | 25mg/L culture |
| Purity | many strong bands on SDS PAGE |
| Notes | |