Refolding Record:
| Protein | |
|---|---|
| Protein Name | Urokinase-type plasminogen activator |
| Abbreviated Name | uPA |
| SCOP Family | Kringle Modules |
| Structure Notes | |
| Organism | Human |
| UniProt Accession | P00749 |
| SCOP Unique ID | n/a |
| Structure Solved | |
| Class | Small Proteins |
| Molecularity | Monomer |
| Construct | |
|---|---|
| Full Length | y |
| Domain | n/a |
| Chimera | n/a |
| Variants | n/a |
| Chain Length | 438 |
| Molecular Weight | 48525.5 |
| Pi | 8.78082 |
| Molecular Weight | 48525.5 |
| Disulphides | Unknown |
| Full Sequence |
MRALLARLLLCVLVVSDSKGSNELHQVPSNCDCLNGGTCVSNKYFSNIHWCNCPKKFGGQ
HCEIDKSKTCYEGNGHFYRGKASTDTMGRPCLPWNSATVLQQTYHAHRSDALQLGLGKHN
YCRNPDNRRRPWCYVQVGLKPLVQECMVHDCADGKKPSSPPEELKFQCGQKTLRPRFKII
GGEFTTIENQPWFAAIYRRHRGGSVTYVCGGSLMSPCWVISATHCFIDYPKKEDYIVYLG
RSRLNSNTQGEMKFEVENLILHKDYSADTLAHHNDIALLKIRSKEGRCAQPSRTIQTICL
PSMYNDPQFGTSCEITGFGKENSTDYLYPEQLKMTVVKLISHRECQQPHYYGSEVTTKML
CAADPQWKTDSCQGDSGGPLVCSLQGRMTLTGIVSWGRGCALKDKPGVYTRVSHFLPWIR
SHTKEENGLAL
|
| Notes | n/a |
| Expression | |
|---|---|
| Report | Orsini G, Brandazza A, Sarmientos P, Molinari A, Lansen J, Cauet G. (1991) Eur J Biochem., 195, 691-697 |
| Project Aim | Structure-Function |
| Fusion | None |
| Protein Expression and Production | Protein recombinantly expressed as and refolded from inclusion bodies. |
| Expression Host | Escherichia coli |
| Expression Strain | B |
| Expression Temp | 37.0 |
| Expression Time | unknown |
| Expression Vector | pFC16 |
| Expression Protocol | unknown |
| Method of Induction | Not Stated |
| Cell Density at Induction | OD = |
| Cell Disruption Method | Sonication |
| Lytic Agent | None |
| Pre-Refolding Purification | None |
| Solubility | insoluble |
| Refolding | |
|---|---|
| Refolding Method | Dilution |
| Wash Buffer | 0.1% Triton X-100, 10mM Tris-HCl, pH 8 |
| Solubilization Buffer | 6M guanidinium chloride, 10mM Tris-HCl, 50mM betamercaptoethanol, pH 8.5 |
| Refolding Buffer | 2.5M urea, 10mM Tris-HCl, 5mM EDTA |
| Pre-Refolding Purification | None |
| Tag Cleaved | no tag |
| Refolding pH | 7.6 |
| Refolding Temperature | 15.0 |
| Protein Concentration | 0.2/mg/ml |
| Refolding Time | 24h |
| Redox Agent | None |
| Redox Agent Concentration | n/a |
| Refolding Protocol | The wash inclusion bodies were solubilised overnight at 4C. The protein was refolded by 20-30 fold dilution with 2.5M urea, 10mM Tris-HCl, 5mM EDTA, pH 7.6. Maximal activity was detected 20-24h after dilution. |
| Refolding Assay | Enzyme activity |
| Refolding Chaperones | None |
| Refolding Additives | None |
| Additives Concentration | NULL |
| Refolding Yield | 15% |
| Purity | |
| Notes | |