Refolding Record:
| Protein | |
|---|---|
| Protein Name | Pyrimidine dimer DNA glycosylase |
| Abbreviated Name | Pyrimidine dimer DNA glyc |
| SCOP Family | Unknown |
| Structure Notes | |
| Organism | Micrococcus luteus |
| UniProt Accession | P94957 |
| SCOP Unique ID | n/a |
| Structure Solved | |
| Class | Unknown |
| Molecularity | Dimer |
| Construct | |
|---|---|
| Full Length | y |
| Domain | n/a |
| Chimera | n/a |
| Variants | n/a |
| Chain Length | 154 |
| Molecular Weight | 17036.4 |
| Pi | 9.37 |
| Molecular Weight | 17036.4 |
| Disulphides | 0 |
| Full Sequence |
MRLWPLHPRHLDRQGLTGAWREALLAQAVLAGRTRGYRDHPQLLRFREHPDPSGAIGAFLSGLEAEATARGYRFDHSRIDRPCPAPDGAVRIPPAGLAPIPATTGQRDLEWRHLCAKLAVRSPAWLEQWSDTPVPDVHPLFTIVPGPVASERAS
|
| Notes | No details on vector given. |
| Expression | |
|---|---|
| Report | Basilico, G., Roger, C. A., Seigelchifer, M., Kerner, N. (2005) J Dermatol Sci, 39, 81-88 |
| Project Aim | Drug Studies |
| Fusion | None |
| Protein Expression and Production | Protein recombinantly expressed as and refolded from inclusion bodies. |
| Expression Host | Escherichia coli |
| Expression Strain | ENV1 |
| Expression Temp | 42.0 |
| Expression Time | not stated |
| Expression Vector | pEX-IFN-alpha |
| Expression Protocol | Cells grown at 28 degC in LB medium containing 50 microg/mL ampicillin with vigorous agitation. |
| Method of Induction | Temperature Shift |
| Cell Density at Induction | OD n/a = n/a |
| Cell Disruption Method | None |
| Lytic Agent | None |
| Pre-Refolding Purification | None |
| Solubility | insoluble |
| Refolding | |
|---|---|
| Refolding Method | Dilution |
| Wash Buffer | 1mM EDTA, 2 M urea, 0.5% Triton X-100 |
| Solubilization Buffer | 8 M urea, 100 mM glycine, pH 10.0, 100mM NaCl, 5 % Beta-mercaptoethanol |
| Refolding Buffer | 100mM glycine, pH 10, 1 M L-alpha-arginine, 100mM NaCl, 100microM CuCl2. |
| Pre-Refolding Purification | None |
| Tag Cleaved | no tag |
| Refolding pH | 10.0 |
| Refolding Temperature | 28.0 |
| Protein Concentration | n/a |
| Refolding Time | 5h |
| Redox Agent | Beta-mercaptoethanol |
| Redox Agent Concentration | 5%,5% |
| Refolding Protocol | Inclusion bodies were washed and solubilized in appropriate buffers. Refoling was preformed by diluting 1:30 in refoldin buffer in air oxidising conditions. |
| Refolding Assay | SDS-PAGE |
| Refolding Chaperones | None |
| Refolding Additives | L-Arginine |
| Additives Concentration | 1 M |
| Refolding Yield | n/a |
| Purity | n/a |
| Notes | n/a |