Refolding Record:
| Protein | |
|---|---|
| Protein Name | Prohibitin |
| Abbreviated Name | Prohibitin |
| SCOP Family | Unknown |
| Structure Notes | |
| Organism | Human |
| UniProt Accession | P35232 |
| SCOP Unique ID | n/a |
| Structure Solved | |
| Class | Unknown |
| Molecularity | Tetramer |
| Construct | |
|---|---|
| Full Length | y |
| Domain | n/a |
| Chimera | n/a |
| Variants | n/a |
| Chain Length | 284 |
| Molecular Weight | 31366.7 |
| Pi | 5.74 |
| Molecular Weight | 31366.7 |
| Disulphides | 0 |
| Full Sequence |
MAAKVFESIGKFGLALAVAGGVVNSALYNVDAGHRAVIFDRFRGVQDIVVGEGTHFLIPWVQKPIIFDCRSRPRNVPVITGSKDLQNVNITLRILFRPVASQLPRIFTSIGEDYDERVLPSITTEILKSVVARFDAGELITQRELVSRQVSDDLTERAATFGLILDDVSLTHLTFGKEFTEAVEAKQVAQQEAERARFVVEKAEQQKKAAIISAEGDSKAAELIANSLATAGDGLIELRKLEAAEDIAYQLSRSRNITYLPAGQSVLLQLPQPWEDPDHHHHHH
|
| Notes | n/a |
| Expression | |
|---|---|
| Report | Bacher, S., Achatz, G., Schmitz, M. L., Lamers, M. C. (2002) Biochimie, 84, 1205-1218 |
| Project Aim | Crystallography,Protein-Protein Interaction Identification |
| Fusion | C-terminal hexahistidine tag |
| Protein Expression and Production | Protein recombinantly expressed as and refolded from inclusion bodies. |
| Expression Host | Escherichia coli |
| Expression Strain | M15 |
| Expression Temp | 37.0 |
| Expression Time | 6h |
| Expression Vector | pQE-60 |
| Expression Protocol | 1 L of LB medium containing ampicillin and kanamycin was inoculated with cells and grown to appropri |
| Method of Induction | IPTG |
| Cell Density at Induction | OD 600 = 0.7 |
| Cell Disruption Method | None |
| Lytic Agent | None |
| Pre-Refolding Purification | None |
| Solubility | insoluble |
| Refolding | |
|---|---|
| Refolding Method | Column refolding: Nickel-chelating chromatography |
| Wash Buffer | n/a |
| Solubilization Buffer | 6 M GuHCl, 0.1 M NaH2PO4, 0.01 M Tris·Cl, pH 8.0 |
| Refolding Buffer | decreasing concentration denaturant 0.1 M NaH2PO4, 0.01 M Tris·Cl, pH 8.0 |
| Pre-Refolding Purification | None |
| Tag Cleaved | no |
| Refolding pH | 8.0 |
| Refolding Temperature | 37.0 |
| Protein Concentration | n/a |
| Refolding Time | n/a |
| Redox Agent | None |
| Redox Agent Concentration | n/a |
| Refolding Protocol | Recombinant prtoteins were purified by the addition of denaturing buffer (Qiagen) and refolded on a Ni-NTA agrose column by sequentail washing with decreasing concentrations of denaturant. Protein was eluted by washing with HBS containing 100mM imidazole, followed by dialysis. |
| Refolding Assay | Surface plasmon resonance binding |
| Refolding Chaperones | None |
| Refolding Additives | None |
| Additives Concentration | n/a |
| Refolding Yield | n/a |
| Purity | n/a |
| Notes | n/a |