Refolding Record:
| Protein | |
|---|---|
| Protein Name | Growth Hormone |
| Abbreviated Name | GH |
| SCOP Family | Long-Chain Cytokines |
| Structure Notes | |
| Organism | Human |
| UniProt Accession | P01241 |
| SCOP Unique ID | n/a |
| Structure Solved | |
| Class | Alpha |
| Molecularity | Monomer |
| Construct | |
|---|---|
| Full Length | y |
| Domain | n/a |
| Chimera | n/a |
| Variants | n/a |
| Chain Length | 217 |
| Molecular Weight | 24847.3 |
| Pi | 5.28701 |
| Molecular Weight | 24847.3 |
| Disulphides | 2 |
| Full Sequence |
MATGSRTSLL LAFGLLCLPW LQEGSAFPTI PLSRLFDNAM LRAHRLHQLA FDTYQEFEEA YIPKEQKYSF LQNPQTSLCF SESIPTPSNR EETQQKSNLE LLRISLLLIQ SWLEPVQFLR SVFANSLVYG ASDSNVYDLL KDLEEGIQTL MGRLEDGSPR TGQIFKQTYS KFDTNSHNDD ALLKNYGLLY CFRKDMDKVE
TFLRIVQCRS VEGSCGF
|
| Notes | n/a |
| Expression | |
|---|---|
| Report | Tchelet A, Vogel T, Helman D, Guy R, Neospouolus C, Goffin V, Djiane J, Gertler A (1997) Mol Cell Endocrinol, 130, 141-152 |
| 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 | BL21 |
| Expression Temp | 42.0 |
| Expression Time | not stated |
| Expression Vector | pTVT715-15 |
| Expression Protocol | no details provided |
| 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 | Dialysis |
| Wash Buffer | n/a |
| Solubilization Buffer | 4.5M urea, 40mM Tris |
| Refolding Buffer | 10mM TrisHCl pH 8.0 |
| Pre-Refolding Purification | None |
| Tag Cleaved | no tag |
| Refolding pH | 8.0 |
| Refolding Temperature | 4.0 |
| Protein Concentration | |
| Refolding Time | |
| Redox Agent | Cysteine |
| Redox Agent Concentration | 1mM |
| Refolding Protocol | A 40g cake of pelleted E.coli cells was suspended in 160ml TrisHCl pH 8.0, 50mM EDTA and sonicated. The inclusion body pellet was solubilized in 300ml solubilization buffer, the pH was increased to 11.3 with NaOH, cysteine was added to 1mM and the solution was stirred for 48h at 4degC, then dialysed against 4 x 10L of refolding buffer. The refolded protein was then purified using a Q-sepharose column followed by gel filtration chromatography on a Superdex 75 column. |
| Refolding Assay | Far-UV Circular Dichroism |
| Refolding Chaperones | None |
| Refolding Additives | None |
| Additives Concentration | |
| Refolding Yield | |
| Purity | |
| Notes | |