Construct Design
Most target constructs are designed based on experience, precedent and downstream application. For difficult targets, we use focused small-scale screens to find suitable boundaries.
Tags
| Affinity | Solubility | Affinity & Solubility | Protease Site |
|---|---|---|---|
| His-Tag HHHHHH | SUMO | MBP | TEV ENLYFQ\S |
| StrepII-Tag WSHPQFEK |  |
GST | Thrombin LVPR\GS |
| Flag-Tag DYKDDDDK | Fc | Enterokinase DDDDK\X |
Plasmids
| E.coli | pET15, pET28, pET41, pMAL, pET-Duet, pET-Blue |
|---|---|
| Insect | pFastBac, pFastBac-Dual |
| Mammalian | pCAGGS |
Cell Lines
| BL21(DE3) | pLysS | Rosetta | Arctic | SF9 - SF21 Hi5 | HEK293, CHO-S, HEK293-6E |
|---|---|---|---|---|---|
| Most Targets in E. coli | Toxic Proteins | Rare Codons | Folding Chaperones | Insect Cell Expression | Mammalian Cell Expression |
Product Production
Purification
We purify all proteins in-house on AKTA FPLC systems. Our standard 3-step protocol includes affinity, ion exchange and size-exclusion chromatography typically yields > 95% purity.
Formulation
Many target proteins are readily stabilized in a standard buffer near physiological pH. If needed, we can screen additional components (ie. glycerol, sucrose, protease inhibitors) to formulate the protein for your downstream applications.
Validation
After purification all protein can be tested with UV spectrophotometry and SDS page gel analysis to ensure a >95% pure product. A Certificate of Analysis (COA) will be included with the protein containing all the testing results and handling instructions.
Protein quality is key for success in structural biology. At SBD we are experts in expressing and purifying “crystallography grade” recombinant protein. We express your target in E. coli or insect cells, purify it on AKTA systems, and formulate it for your downstream applications. Available as stand-alone service or combine it with crystal structure determination and activity assays.
Contact us for a consultation and quote for your project.