Refolding Record:
| Protein | |
|---|---|
| Protein Name | Streptavidin |
| Abbreviated Name | SCD |
| SCOP Family | Avidin/streptavidin |
| Structure Notes | |
| Organism | Streptomyces avidinii |
| UniProt Accession | P22629 |
| SCOP Unique ID | n/a |
| Structure Solved | |
| Class | Beta |
| Molecularity | Dimer |
| Construct | |
|---|---|
| Full Length | n |
| Domain | n/a |
| Chimera | n/a |
| Variants | Single Chained Dimeric |
| Chain Length | 252 |
| Molecular Weight | 26259.5 |
| Pi | 6.11 |
| Molecular Weight | 26259.5 |
| Disulphides | 0 |
| Full Sequence |
MAEAGITGTWYNQLGSTFIVTAGADGALTGTYESAVGNAESRYVLTGRYDSAPATDGSGTALGWTVAWKNNYRNAHSATTWSGQYVGGAEARINTQWLLTSGTTEANAWKSTLVGHDTFTKVKPSAASGGGSSAEAGITGTWYNQLGSTFIVTAGADGALTGTYESAVGNAESRYVLTGRYDSAPATDGSGTALGWTVAWKNNYRNAHSATTWSGQYVGGAEARINTQWLLTSGTGSGFFLAAALEHHHHHH
|
| Notes | n/a |
| Expression | |
|---|---|
| Report | Aslan, F. M., Yu, Y., Mohr, S., Cantor, C. R. (2005) Proc. Natl. Acad. Sci. USA, 102, 8507-8512 |
| Project Aim | Structure-Function |
| Fusion | C-terminal hexahistidine tag |
| Protein Expression and Production | Protein recombinantly expressed as and refolded from inclusion bodies. |
| Expression Host | Escherichia coli |
| Expression Strain | BL21(DE3)Gold-pLysS |
| Expression Temp | 37.0 |
| Expression Time | 2 |
| Expression Vector | pET22b(+) |
| Expression Protocol | Cells were washed with 200ml buffer (10mM TrisHCl, pH 8, 100m M NaCL, 1 mM EDTA) and resuspended in |
| Method of Induction | IPTG |
| Cell Density at Induction | OD n/a = n/a |
| Cell Disruption Method | Sonication |
| Lytic Agent | None |
| Pre-Refolding Purification | None |
| Solubility | insoluble |
| Refolding | |
|---|---|
| Refolding Method | Dilution |
| Wash Buffer | n/a |
| Solubilization Buffer | 6 M Gu.HCl, 50 mM TrisHCL, pH 7.5 (or 7 M Gu.HCl, pH 1.5 (5.0 ml/litre culture)) |
| Refolding Buffer | 50mM TrisHCl, 150mM NaCl, 5 mM EDTA, 0.1 mM PMSF, pH 7.5 |
| Pre-Refolding Purification | None |
| Tag Cleaved | no |
| Refolding pH | 7.5 |
| Refolding Temperature | 4.0 |
| Protein Concentration | n/a |
| Refolding Time | n/a |
| Redox Agent | None |
| Redox Agent Concentration | n/a |
| Refolding Protocol | Solubilisation buffer was added to the homogenized solution and was allowed to equilibrate for several hours at 4degC before centrifugation (18 000g, 10min, 4degC). Solubilized protein was diluted dropwise (1 drop per sec) into refolding buffer and left to equilibrate overnight. |
| Refolding Assay | Ligand Binding |
| Refolding Chaperones | None |
| Refolding Additives | None |
| Additives Concentration | n/a |
| Refolding Yield | n/a |
| Purity | n/a |
| Notes | n/a |