Refolding Record:
| Protein | |
|---|---|
| Protein Name | CDC10-dependent transcription 1 |
| Abbreviated Name | CDT1 |
| SCOP Family | Coiled-coil |
| Structure Notes | |
| Organism | Xenopus laevis |
| UniProt Accession | Q9I9A7 |
| SCOP Unique ID | n/a |
| Structure Solved | |
| Class | Unknown |
| Molecularity | Unknown |
| Construct | |
|---|---|
| Full Length | y |
| Domain | n/a |
| Chimera | n/a |
| Variants | n/a |
| Chain Length | 630 |
| Molecular Weight | 69853.5 |
| Pi | 9.8124 |
| Molecular Weight | 69853.5 |
| Disulphides | Unknown |
| Full Sequence |
MADMSQMRVTDFFSQSKRGTAAQNSKGRKVEAVLETRRAVTRSRAASVKAEEFLKAPCTP
ERASPTVSQCIGPSSKKRTRQDSDSEPLRTQQRQGKSARKKLKLPEGEHGGSVQQQLFSP
CNKVALEHVTPSSLGKKIKDMVNVSLSPKFNELARNPTTPETKSPAKENLLELKSRLQRI
QELAQKVNLPAASSEGKVTITDLKARLKRAQELDTKIRAKAEKTETQAIDLTEQPAQESE
KAPAYQRFHNLAQDAAPGLTLPYKYKVLAEMFRSMDTIVGMLFNRSETITFSKVKQGVQD
MMRKQFEQRNVGQIKTVYPNAYKYRQEKNIPTFKDGVKKTDYQLTIEPLVAEGDMLSGRP
HLSASRLLERKQLFHRSLTSIVKQHHRVFLTSLNPPMLVPDDKLTRWHPRFNVDEVLDVT
PAELPLPPQVERLTTAQEVLSKARGLITPKMEKALANLALKTAENAGETKNVSTEETKST
ATTSTSTALKGVSQSLLERIRAKEAQKLQAIMTRRPQQEERLLMMSRLPELARILRNVFV
AEKKPALTLEVTCSRVIASCRSSMSPGEMEKHLALLSEILPDWLSIHPVRKDTYYKLNQS
MDLNLILERLAKKTKEEESL
|
| Notes | n/a |
| Expression | |
|---|---|
| Report | MAIORANO, D., Moreau, J., Mechali, M. (2000) Nature, 404, 622-625 |
| Project Aim | Functional Studies |
| Fusion | N-terminal hexahistidine tag |
| Protein Expression and Production | Protein recombinantly expressed as and refolded from inclusion bodies. |
| Expression Host | Escherichia coli |
| Expression Strain | BL21 |
| Expression Temp | 37.0 |
| Expression Time | 3 hours |
| Expression Vector | pRSET |
| 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 | 100 mM NaH2PO4, 10 mM TrisHCl, 8 M Urea, pH 6.3 |
| Solubilization Buffer | 100 mM NaH2PO4, 10 mM TrisHCl, 8 M Urea, pH 8.0 |
| Refolding Buffer | 50 mM Hepes, 0.2M NaCl, 1 mM DTT, 1 M NDSB201 |
| Pre-Refolding Purification | None |
| Tag Cleaved | no |
| Refolding pH | 7.5 |
| Refolding Temperature | 4.0 |
| Protein Concentration | 0.1 mg/ml |
| Refolding Time | 1 hour |
| Redox Agent | DTT |
| Redox Agent Concentration | n/a |
| Refolding Protocol | Dilute your protein 10-fold into ice-cold refolding buffer as quickly as possible so that the final protein concentration will not exceed 0.1-0.05 mg/ml. For small volumes (1-10 ml) the protein can be added directly into the refolding buffer in a 15 ml Falcon tube while vortexing. Keep vortexing for 30 seconds after addition. For larger volumes, the protein can be added into a becker containing the refolding buffer while vigourously stirring. Keep stirring for 2 minutes after addition. Leave on ice 1 hour. The protein can be now dyalized and concentrated by centrifugation through a Microcon-30 device (Millipore) at 4 °C. |
| Refolding Assay | Bioactivity |
| Refolding Chaperones | None |
| Refolding Additives | None |
| Additives Concentration | NULL |
| Refolding Yield | 1 mg/ml |
| Purity | 99 % |
| Notes | |