Refolding Record:
| Protein | |
|---|---|
| Protein Name | CREB-binding protein |
| Abbreviated Name | CBP |
| SCOP Family | TAZ domain |
| Structure Notes | |
| Organism | Mouse |
| UniProt Accession | P45481 |
| SCOP Unique ID | n/a |
| Structure Solved | |
| Class | Small Proteins |
| Molecularity | Monomer |
| Construct | |
|---|---|
| Full Length | n |
| Domain | TAZ2 domain (aa.1764-1850) |
| Chimera | n/a |
| Variants | n/a |
| Chain Length | 88 |
| Molecular Weight | 9982.9 |
| Pi | 9.55 |
| Molecular Weight | 9982.9 |
| Disulphides | 0 |
| Full Sequence |
MSPQESRR LSIQRCIQSL VHACQCRNAN CSLPSCQKMK RVVQHTKGCK RKTNGGCPVC KQLIALCCYH AKHCQENKCP VPFCLNIKHK
|
| Notes | Zinc-binding domain |
| Expression | |
|---|---|
| Report | DeGuzman RN, Liu HY, Martinez-Yamout M, Dyson HJ, Wright PE (2000) J Mol Biol, 303, 243-253 |
| Project Aim | NMR |
| Fusion | None |
| Protein Expression and Production | Protein recombinantly expressed as and refolded from inclusion bodies. |
| Expression Host | Escherichia coli |
| Expression Strain | BL21(DE3) |
| Expression Temp | 37.0 |
| Expression Time | not stated |
| Expression Vector | pET3a |
| Expression Protocol | not stated |
| Method of Induction | Not Stated |
| Cell Density at Induction | OD n/a = n/a |
| Cell Disruption Method | Not stated |
| Lytic Agent | None |
| Pre-Refolding Purification | HPLC |
| Solubility | insoluble |
| Refolding | |
|---|---|
| Refolding Method | Dilution |
| Wash Buffer | n/a |
| Solubilization Buffer | 6M GdnHCl |
| Refolding Buffer | 20mM sodium acetate pH 8.5, 20mM DTT, 3-4 molar equilvalent of ZnCl2 |
| Pre-Refolding Purification | HPLC |
| Tag Cleaved | no tag |
| Refolding pH | 8.5 |
| Refolding Temperature | 4.0 |
| Protein Concentration | 200microM |
| Refolding Time | n/a |
| Redox Agent | DTT |
| Redox Agent Concentration | 20mM |
| Refolding Protocol | Inclusion bodies were dissolved in 6M GdnHCl and purified by HPLC. The purified protein was reconstituted with Zn2+ by being dissolved in refolding buffer to a final concentration of about 200microM. The pH was decreased to 5.8 with 1M HCl, excess DTT and ZnCl2 were removed and the protein was concentrated to 1.5mM |
| Refolding Assay | Far-UV Circular Dichroism,15N -1H chemical shifts (ppm) |
| Refolding Chaperones | None |
| Refolding Additives | None |
| Additives Concentration | n/a |
| Refolding Yield | n/a |
| Purity | n/a |
| Notes | n/a |