Refolding Record:
| Protein | |
|---|---|
| Protein Name | Lumazine synthase 2 |
| Abbreviated Name | Lumazine synthase 2 |
| SCOP Family | Lumazine synthase |
| Structure Notes | |
| Organism | Brucella abortus |
| UniProt Accession | P61711 |
| SCOP Unique ID | n/a |
| Structure Solved | |
| Class | Alpha/Beta |
| Molecularity | Pentamer |
| Construct | |
|---|---|
| Full Length | n |
| Domain | aa.1-145 |
| Chimera | n/a |
| Variants | n/a |
| Chain Length | 145 |
| Molecular Weight | 15932.2 |
| Pi | 6.39358 |
| Molecular Weight | 15932.2 |
| Disulphides | 0 |
| Full Sequence |
MNQSCPNKTS FKIAFIQARW HADIVDEARK SFVAELAAKT GGSVEVEIFD VPGAYEIPLH AKTLARTGRY AAIVGAAFVI DGGIYRHDFV ATAVINGMMQ VQLETEVPVL SVVLTPHHFH ESKEHHDFFH AHFKVKGVEA AHAAL
|
| Notes | n/a |
| Expression | |
|---|---|
| Report | Goldbaum FA, Polikarpov I, CauerhffAA, Velikovsky CA, Braden BC, Poljak RJ (1998) Mol Cell Endocrinol, 123, 175-178 |
| Project Aim | Crystallography |
| Fusion | None |
| Protein Expression and Production | Protein recombinantly expressed as and refolded from inclusion bodies. |
| Expression Host | Escherichia coli |
| Expression Strain | BL21(DE3) |
| Expression Temp | 37.0 |
| Expression Time | 4 |
| Expression Vector | pET11b |
| Expression Protocol | Cultures were incubated in LB medium at 37degC, expression was induced with 1mM IPTG and cells were grown for 4h. |
| Method of Induction | IPTG |
| Cell Density at Induction | OD = |
| Cell Disruption Method | Sonication |
| Lytic Agent | None |
| Pre-Refolding Purification | None |
| Solubility | insoluble |
| Refolding | |
|---|---|
| Refolding Method | Dilution |
| Wash Buffer | n/a |
| Solubilization Buffer | 50mM Tris, 5mM EDTA, 8M urea, pH 8.0 |
| Refolding Buffer | 150mM NaCl, 10mM NaPhosphate pH 7.4, 1mM DTT |
| Pre-Refolding Purification | None |
| Tag Cleaved | no tag |
| Refolding pH | 7.4 |
| Refolding Temperature | 4.0 |
| Protein Concentration | |
| Refolding Time | 3 days |
| Redox Agent | DTT |
| Redox Agent Concentration | 1mM |
| Refolding Protocol | The cells were sonicated and inclusion bodies werer dissolved in solubilization buffer at room temperature with overnight agitation. the protein was refolded by dialysis against refolding buffer during 3 days with several changes of buffer. The protein was then purified using a MonoQ column. |
| Refolding Assay | Crystallography |
| Refolding Chaperones | None |
| Refolding Additives | None |
| Additives Concentration | |
| Refolding Yield | |
| Purity | |
| Notes | |