Refolding Record:
| Protein | |
|---|---|
| Protein Name | Lupanine hydroxylase |
| Abbreviated Name | Lupanine hydroxylase |
| SCOP Family | Unknown |
| Structure Notes | |
| Organism | Pseudomonas sp. |
| UniProt Accession | Q934G0 |
| SCOP Unique ID | n/a |
| Structure Solved | |
| Class | Unknown |
| Molecularity | Monomer |
| Construct | |
|---|---|
| Full Length | y |
| Domain | n/a |
| Chimera | n/a |
| Variants | n/a |
| Chain Length | 680 |
| Molecular Weight | 72256.9 |
| Pi | 5.7541 |
| Molecular Weight | 72256.9 |
| Disulphides | Unknown |
| Full Sequence |
NEKDGSAVTSGNWSLLGGGNEQHYFSALKDVNKS
NVKNLGLSWFTDMEAGDGLVGNPLVADGVIYQGGPPGKIYANDLKTGKNLWTYTPEVQYD
KDTSWTGFWGTHVNRGLAVDDDNVYIGSYCKLLAVSRTTHKLTWSSQSCDPKKMQAITGA
PRVGGGKVFIGNASGDFGGDRGHLDAFDAKTGKHLWRFYTMPGDPSKPFENDLLAKASKT
WGTDYWKYTKGGVSPWDAITYDEASDTLYFGTDGPSPWSPAQRAPDAGDELFSHSIIAVD
ASTGAYKWHFQTVQNDGSNMSATMHIMLADLPVEGVSKRVVMTAPKNGYFYVLDASTGKF
ISADHYVPVNWTKGLDPKTGRPIPSNEANYWERPGEMTIPLPGDVGGHNWEAMAYNPELR
TVYIPSTLVPVTVVASKDTGELDLDYYYGMRPDATIKTQGDLVAWDPLLQKEKWRAKRSL
PVNGGVLATAGGLVFQGTGDGHFEAFDANTGEKLWSFHVGGSILAAPTTVEVDGDQYLIV
ASGNGGASGMRGIPRLMNNLQSQGPARLLAFRLGGKTELPITSTPDFPKPQYPKPTSAMA
ESGRHIFNANACGACHGFNAEGSTPGLPDLRRSDKLDLAVMKSIVIDGAFKPLGMPGHPH
ISDADLQALQAFILQKAWTAYDTQQTLKTSDTGAQ
|
| Notes | n/a |
| Expression | |
|---|---|
| Report | Hopper DJ, Kaderbhai MA, Marriott SA, Young M, Rogozinski J. (2002) Biochem J., 367, 483-489 |
| Project Aim | Functional Studies |
| Fusion | None |
| Protein Expression and Production | Protein recombinantly expressed as and refolded from inclusion bodies. |
| Expression Host | Escherichia coli |
| Expression Strain | TB1 |
| Expression Temp | 37.0 |
| Expression Time | 12h |
| Expression Vector | pLiQ |
| 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 | unknown |
| Solubilization Buffer | 8M urea |
| Refolding Buffer | 10mM Tris-HCl, 1mM EDTA, 0.13mM PMSF, 0.2mM 1,10-phananthroline |
| Pre-Refolding Purification | None |
| Tag Cleaved | no tag |
| Refolding pH | 8.0 |
| Refolding Temperature | 20.0 |
| Protein Concentration | |
| Refolding Time | 30 |
| Redox Agent | None |
| Redox Agent Concentration | n/a |
| Refolding Protocol | The solubilised protein was refolded by dilution in 1.5L of 10mM Tris-HCl, 1mM EDTA, 0.13mM PMSF, 0.2mM 1,10-phananthroline. In order to activate the enzyme, CaCl2 and pyrroloquinoline quinone were supplemented in the soltuion of refolded protein. |
| Refolding Assay | Enzyme activity |
| Refolding Chaperones | None |
| Refolding Additives | None |
| Additives Concentration | NULL |
| Refolding Yield | |
| Purity | |
| Notes | |