Refolding Record:
| Protein | |
|---|---|
| Protein Name | Bikunin Placental |
| Abbreviated Name | HAI-2 |
| SCOP Family | Unknown |
| Structure Notes | |
| Organism | Human |
| UniProt Accession | O43291 |
| 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 | 225 |
| Molecular Weight | 25415.7 |
| Pi | 8.26 |
| Molecular Weight | 25415.7 |
| Disulphides | 6 |
| Full Sequence |
ADRERSIHDFCLVSKVVGRCRASMPRWWYNVTDGSCQLFVYGGCDGNSNNYLTKEECLKKCATVTENATGDLATSRNAADSSVPSAPRRQDSEDHSSDMFNYEEYCTANAVTGPCRASFPRWYFDVERNSCNNFIYGGCRGNKNSYRSEEACMLRCFRQQENPPLPLGSKVVVLAGLFVMVLILFLGASMVYLIRVARRNQERALRTVWSSGDDKEQLVKNTYVL
|
| Notes | n/a |
| Expression | |
|---|---|
| Report | Seefeldt MB, Ouyang J, Froland WA, Carpenter JF, Randolph TW. (2004) Protein Science, 13, 2639-2650 |
| Project Aim | Protein refolding |
| 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 | None |
| Solubility | |
| Refolding | |
|---|---|
| Refolding Method | Dilution |
| Wash Buffer | n/a |
| Solubilization Buffer | 6M guanidine HCl and 12 mM DTT |
| Refolding Buffer | 50 mM Tris (pH 8.0), 157 mM NaCl, 4 mM GSSG, 2 mM DTT, and 1 M guanidine HCl |
| Pre-Refolding Purification | None |
| Tag Cleaved | no tag |
| Refolding pH | 8.0 |
| Refolding Temperature | 25.0 |
| Protein Concentration | 0.4 mg/mL |
| Refolding Time | 8 h |
| Redox Agent | GSSG/DTT |
| Redox Agent Concentration | 4/2 mM |
| Refolding Protocol | Dilution refolding was conducted by rapidly diluting solutions containing 50 mM Tris (pH 8.0), 157 mM NaCl, 12 mM DTT, and 6 M guanidine HCl to a final solution containing 50 mM Tris (pH 8.0), 157 mM NaCl, 4 mM GSSG, 2 mM DTT, and 1 M guanidine HCl at a protein concentration of 0.4 mg/mL. After dilution, the samples were incubated at 25°C for 8 h (Clark et al. 1998). The samples were then frozen and stored until analysis by SEC-HPLC, RP-HPLC, GEMMA, and activity assays. |
| Refolding Assay | RP-HPLC,Activity assay,SEC-HPLC |
| Refolding Chaperones | None |
| Refolding Additives | None |
| Additives Concentration | n/a |
| Refolding Yield | 55% (±6%) |
| Purity | n/a |
| Notes | n/a |