Refolding Record:
| Protein | |
|---|---|
| Protein Name | CapZ beta-1 subunit |
| Abbreviated Name | CapZ beta-1 subunit |
| SCOP Family | Capz beta-1 subunit |
| Structure Notes | |
| Organism | Chicken (Gallus gallus) |
| UniProt Accession | P14315 |
| SCOP Unique ID | n/a |
| Structure Solved | |
| Class | Multi-domain proteins (alpha and beta) |
| Molecularity | Heterodimer |
| Construct | |
|---|---|
| Full Length | y |
| Domain | n/a |
| Chimera | n/a |
| Variants | n/a |
| Chain Length | 281 |
| Molecular Weight | 31364.5 |
| Pi | 5.36156 |
| Molecular Weight | 31364.5 |
| Disulphides | 0 |
| Full Sequence |
MSDQQLDCALDLMRRLPPQQIEKNLSDLIDLVPSLCEDLLSSVDQPLKIARDKVVGKDYL
LCDYNRDGDSYRSPWSNKYDPPLEDGAMPSARLRKLEVEANNAFDQYRDLYFEGGVSSVY
LWDLDHGFAGVILIKKAGDGSKKIKGCWDSIHVVEVQEKSSGRTAHYKLTSTVMLWLQTN
KTGSGTMNLGGSLTRQMEKDETVSDSSPHIANIGRLVEDMENKIRSTLNEIYFGKTKDIV
NGLRSIDAIPDNQKYKQLQRELSQVLTQRQIYIQPDN
|
| Notes | n/a |
| Expression | |
|---|---|
| Report | Remmert K, Vullhorst D, Hinssen H. (2000) Protein Expression and Purification, 18, 11-19 |
| Project Aim | Undefined |
| Fusion | None |
| Protein Expression and Production | Protein recombinantly expressed as and refolded from inclusion bodies. |
| Expression Host | Escherichia coli |
| Expression Strain | None |
| Expression Temp | 37.0 |
| Expression Time | 4h |
| Expression Vector | pQE60 |
| Expression Protocol | unknown |
| Method of Induction | Not Stated |
| Cell Density at Induction | OD = |
| Cell Disruption Method | Sonication |
| Lytic Agent | None |
| Pre-Refolding Purification | Ion-exchange chromatography |
| Solubility | insoluble |
| Refolding | |
|---|---|
| Refolding Method | Dilution |
| Wash Buffer | 0.1% TritonX-100, 1 mM PMSF, 0.2 mM DTT, 1 mM EGTA, and 10mM Tris, pH 7.5 |
| Solubilization Buffer | 8 M urea, 1 mM EGTA, 0.2 mM DTT, 1 mM PMSF, and 20 mM imidazole, pH 7.4 |
| Refolding Buffer | 20% glycerol, 5 mM EGTA, 2 mMDTT, 1 mMPMSF, and 100mM Tris-HCl, 60mM urea |
| Pre-Refolding Purification | Ion-exchange chromatography |
| Tag Cleaved | no tag |
| Refolding pH | 7.4 |
| Refolding Temperature | 15.0 |
| Protein Concentration | 0.01-0.02 mg/ml |
| Refolding Time | 24h |
| Redox Agent | None |
| Redox Agent Concentration | n/a |
| Refolding Protocol | The CapZ alpha-2 and beta-1 subunits were refolded simultaneously at equimolar concentrations in the same solution to yield heterodimeric product. |
| Refolding Assay | Actin Binding Activity |
| Refolding Chaperones | None |
| Refolding Additives | None |
| Additives Concentration | NULL |
| Refolding Yield | 50% |
| Purity | 50% |
| Notes | |