Refolding Record:
Protein | |
---|---|
Protein Name | Hen egg- white lysozyme |
Abbreviated Name | HEWL |
SCOP Family | C-type Lysozyme |
Structure Notes | |
Organism | Hen |
UniProt Accession | P00698 |
SCOP Unique ID | n/a |
Structure Solved | |
Class | Alpha+Beta |
Molecularity | Unknown |
Construct | |
---|---|
Full Length | y |
Domain | n/a |
Chimera | n/a |
Variants | n/a |
Chain Length | 130 |
Molecular Weight | 14313.1 |
Pi | 9.32 |
Molecular Weight | 14313.1 |
Disulphides | 4 |
Full Sequence |
KVFGRCELAAAMKRHGLDNYRGYSLGNWVCAAKFESNFNTQATNRNTDGSTDYGILQINSRWWCNDGRTPGSRNLCNIPCSALLSSDITASVNCAKKIVSDG
NGMNAWVAWRNRCKGTDVQAWIRGCRL
|
Notes | n/a |
Expression | |
---|---|
Report | Rozema D, Gellman SH (1996) Biochemistry, 35, 15760-15771 |
Project Aim | Protein refolding |
Fusion | None |
Protein Expression and Production | Protein expressed and purified in native conformation prior to denaturation and refolding. |
Expression Host | None |
Expression Strain | None |
Expression Temp | 0.0 |
Expression Time | 0 |
Expression Vector | |
Expression Protocol | |
Method of Induction | Not Stated |
Cell Density at Induction | OD = |
Cell Disruption Method | None |
Lytic Agent | None |
Pre-Refolding Purification | None |
Solubility |
Refolding | |
---|---|
Refolding Method | Dilution |
Wash Buffer | 8.7 M GdmCl, 143 mM Tris sulfate, and 43 mM DTT |
Solubilization Buffer | 0.071 mg/mL lysozyme, 8.6 mM GdmCl, 0.04 mM DTT, 0.071 mM CuSO4, 1.43 mM EDTA, and 0 or 5.7 mM CTAB |
Refolding Buffer | 0.05 mg/mL lysozyme, 6.0 mM GdmCl, 0.03 mM DTT, 0.05 mM CuSO4, 1.0 mM EDTA, 0 or 4.0 mM CTAB, and 4.8 mM -cyclodextrin |
Pre-Refolding Purification | None |
Tag Cleaved | no tag |
Refolding pH | 8.5 |
Refolding Temperature | 25.0 |
Protein Concentration | n/a |
Refolding Time | 40 |
Redox Agent | DTT |
Redox Agent Concentration | 0.03 mM |
Refolding Protocol | Copper sulfate-catalyzed oxidation of denatured-reduced lysozyme was performed by dilution of 50 mg/mL of denatured-reduced lysozyme in 6 M GdmCl and 30 mM DTT to 0.071 mg/mL lysozyme, 8.6 mM GdmCl, 0.04 mM DTT, 0.071 mM CuSO4, 1.43 mM EDTA, and 0 or 5.7 mM CTAB. The solutions were allowed to sit for various periods of time, and then -cyclodextrin was added to make a solution of 0.05 mg/mL lysozyme, 6.0 mM GdmCl, 0.03 mM DTT, 0.05 mM CuSO4, 1.0 mM EDTA, 0 or 4.0 mM CTAB, and 4.8 mM B-cyclodextrin. After sitting 40 h, the solutions were assayed for enzymatic activity. |
Refolding Assay | Enzyme activity |
Refolding Chaperones | None |
Refolding Additives | β-cyclodextrin |
Additives Concentration | 4.8 mM |
Refolding Yield | n/a |
Purity | n/a |
Notes | Please note temperature is not specified assumed it is in room temperature |