Lefty B, Recombinant, Human (Left-Right Determination, Factor B, LEFTB, LEFTY1) (Human 293 Cells)
|Molecular Biology||Storage: -20°CShipping: Blue Ice|
Human Cell Expression Cytokines:
Expression of proteins in our Human 293-cell expression system is a cost-effective and scalable production method for high-authenticity, recombinant human proteins.
Functional activities of human proteins depend on proper folding, phosphorylation, disulfide bridge formation, and proteolytic processing or appropriate glycosylation. Ideally, recombinant human proteins are produced in human cells that are more authentic in terms of both physical properties and biochemical functions. However, the current process of human cell expression requires a large quantity of DNA and medium supplemented with bovine serum. Hence, it is often a daunting task to produce sufficient amount of high quality proteins for drug discovery or diagnostic development (1-100mg) at acceptable cost. United States Biological is now supplying a high yield, cost-effective and versatile system to produce active recombinant human proteins in a human cell line to meet the special needs of the broad scientific community and drug discovery industry. In this system, we have broken key bottleneck steps to increase protein production and reduce cost, including
• Engineered human cell line adapted to suspension for growth in serum-free and chemically defined medium
• Proprietary expression vectors driven by strong promoters and signal peptides
• Large scale transient expression with small quantity of DNA
• Rapid creation of stable human cell lines and adaptation to serum-free medium
• Short time to scale up production
• Efficient tag or tag-free purification
Using this expression system, we have produced a large number of highly active human cytokines, kinases and phosphatases. Our technology is particularly suitable for the production of high-authenticity human proteins in medium scale as research and diagnostic reagents, drug screening targets, and therapeutic proteins for preclinical evaluation. Key differences of recombinant human protein production in different systems are illustrated below:
Expression system E. coli Insect cell CHO cell Human cell
Protein folding + ++ +++ ++++
Phosphorylation - ++ +++ ++++
Proteolytic processing - + +++ ++++
Glycosylation - Poor Not human-like Authentic
LEFTY-B is expressed in human 293 cells as a glycosylated monomer with an apparent molecular mass of 40kD. Production in human 293 cells offers authentic glycosylation. Glycosylation contributes to stability in cell growth media and other applications.
38kD monomer, glycosylated
During vertebrate embryogenesis, a left-right axis is established. Secreted growth factors of the TGF-beta family, including gene products derived from nodal, lefty-1 and lefty-2, play crucial roles in establishing left-right asymmetries. TGF-beta (Transforming growth factor-beta) is a pleiotropic cytokine that regulates growth and differentiation of diverse types of cells. TGF-beta actions are directed by ligand-induced activation of TGF-beta receptors. Complexes formed move into the nucleus, where they act as components of a transcriptional complex. Lefty, a novel member of the TGF-beta superfamily, inhibits TGF-beta signaling. Lefty acts to inhibit phosphorylation of Smad2 following activation of the TGF-beta receptor. Lefty also inhibits events downstream from R-Smad phosphorylation. Lefty provides a repressed state of TGF-beta-responsive genes The Lefty family is comprised of Lefty 1 and Lefty 2 in mouse, and Lefty A and Lefty B in humans. Members of the TGF-beta superfamily require processing for their activation. Cleavage is therefore an essential step for Lefty activation. Lefty is synthesized as a large inactive precursor (42kD) that must be endoproteolytically processed to release the bioactive polypeptide (28kD and 34kD forms). The 28kD form induces MAPK activity.
Recombinant Human LEFTY-B Expressed in Human 293 Cells.
The specific activity was determined by dose dependent ability to inhibit BMP-4 (6.5ng/ml) induction of alkaline phosphatase production in the MC-3T3-E1 cell line. (Mouse chondrogenic cell line).
Storage and Stability:
Lyophilized powder may be stored at -20°C. Stable for 12 months at -20°C. Reconstitute with sterile ddH2O, 0.1% endotoxin-free recombinant HSA. Aliquot and store at -20°C. Reconstituted product is stable for 6 months at 4 °C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.
Source: HEK 293 cells
Purity: 90% as determined by SDS-PAGE and HPLC. Endotoxin: 1EU/ug
Form: Supplied as a lyophilized powder from 10mM Tris-HCl, pH 7.4, 50mM sodium chloride, 0.5% CHAPS. Reconstitute with sterile ddH2O, 0.1% endotoxin-free recombinant HSA.
Important Note: This product as supplied is intended for research use only, not for use in human, therapeutic or diagnostic applications without the expressed written authorization of United States Biological.