Refolding Record:
| Protein | |
|---|---|
| Protein Name | Lysozyme |
| Abbreviated Name | Lysozyme |
| SCOP Family | C-type Lysozyme |
| Structure Notes | |
| Organism | Chicken (Gallus gallus) |
| UniProt Accession | Q6LEL2 |
| 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 | 45 |
| Molecular Weight | 4953.0 |
| Pi | 9.69 |
| Molecular Weight | 4953.0 |
| Disulphides | Unknown |
| Full Sequence |
MRSLLILVLCFLPLAALGKVFGRCELAAAMKRHGLDNYRGYSLGN
|
| Notes | n/a |
| Expression | |
|---|---|
| Report | Lanckriet H, Middelberg AP. (2004) Biotechnol Prog, 20, 1861-1866 |
| 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/ chaperone-assisted refolding |
| Wash Buffer | n/a |
| Solubilization Buffer | 0.1 M Tris, 1 mM EDTA, 6 M urea and 32 mM DTT, pH 8.1 |
| Refolding Buffer | 0.1 M Tris, 1 mM EDTA, 3 mM cysteine and 0.3 mM cystine, pH 8.1, 0-0.36 M urea and 0-15.4 mM B-CD |
| Pre-Refolding Purification | None |
| Tag Cleaved | no tag |
| Refolding pH | 8.1 |
| Refolding Temperature | 25.0 |
| Protein Concentration | n/a |
| Refolding Time | 24 h |
| Redox Agent | Cysteine/Cystine |
| Redox Agent Concentration | 3/0.3 mM |
| Refolding Protocol | Refolding. One-Step Batch Dilution Refolding. Refolding buffer (0.1 M Tris, 1 mM EDTA, 3 mM cysteine and 0.3 mM cystine, pH 8.1) was used for all refolding experiments. In some cases the buffer also contained 0-0.36 M urea and 0-15.4 mM -CD. Lysozyme refolding was initiated through a 50-fold dilution of denatured lysozyme with refolding buffer. This procedure was executed for denatured lysozyme solutions with and without CTAB. The samples were left to refold for 24 h at room temperature before analysis. Purification. A sample of 500 L of batch refolded lysozyme solution was loaded on a prepacked 25-mL HiTrap desalting column (Amersham Biosciences, Bucks, U.K.) having an inner diameter of 1.6 cm. The column was previously equilibrated with the running buffer (0.1 M Tris, 1 mM EDTA, pH 8.1). The sample was eluted with the same buffer at 1 mL/min. |
| Refolding Assay | Unspecified |
| Refolding Chaperones | None |
| Refolding Additives | β-cyclodextrin |
| Additives Concentration | 0-15.4 mM |
| Refolding Yield | 53% |
| Purity | n/a |
| Notes | n/a |