Recombinant Human TNF RI/TNFRSF1A Protein Summary
| Description |
A biologically active protein to TNFRSF1A. |
| Specificity |
TNF RI-Fc HCX Chimera migrates as a band between 50 and 60 kDa in SDSPAGE due to post-translation modifications, in particular glycosylation. This compares with the unmodified TNF RI-Fc Chimera that has a predicted molecular mass of 48.0kDa. TNF RI-Fc HCX Chimera has N-linked and possibly O-linked oligosaccharides. |
Preparation Method |
A DNA sequence encoding the signal peptide and extracellular domain of human TNF Receptor 1 (TNFRI) (aa 1-209) was fused to the Fc region of human IgG1 (aa 93-330). The chimeric protein was expressed in modified human 293 cells. |
| Details of Functionality |
The ND50 of TNF RI-Fc HCX Chimera is typically 15-30 ng/ml as measured by its ability to neutralize TNF-a mediated cytoxicity in murine WEHI 164 cells in the presence of
actinomycin D. |
| Protein/Peptide Type |
Biologically Active Protein |
| Gene |
TNFRSF1A |
Applications/Dilutions
| Dilutions |
- Block/Neutralize
- Functional
- Western Blot
|
| Application Notes |
This protein is functionally active and can be used for Blocking and Neutralizing. It can also be used for Western Blot. TNF RI-Fc HCX Chimera separates into a number of isoforms with a pI between 6.5 and 8.6 in 2D PAGE due to post-translational modifications, in particular glycosylation.
This compares with the unmodified TNF RI-Fc Chimera that has a predicted pI of 7.69. |
Reactivity Notes
Packaging, Storage & Formulations
| Storage |
Store at -80C. Avoid freeze-thaw cycles. |
| Concentration |
LYOPH |
| Reconstitution Instructions |
Reconstitute with 0.5 ml sterilized PBS. After reconstitution the buffer solution will contain 1% human serum albumin and 10% trehalose already present in the vial. |
Alternate Names for Recombinant Human TNF RI/TNFRSF1A Protein
Background
Tumor necrosis factor alpha (TNF-a ; TNF alpha) and the structurally related lymphotoxin alpha are pro-inflammatory cytokines produced by a variety of immune and non-immune cells in response to multiple stimuli. These stimuli can include bacteria and bacterial products such as LPS, viral infections, T-cell receptor activation and cross-linking of surface immunoglobulin (Ig) on B cells. Expression of TNF-alpha has been detected in macrophages/monocytes, B-cells, T-cells, basophils, eosinophils, glial cells and mast cells. The effects of TNF-a are mediated through the TNF receptors, TNF RI (TNF R1) and TNF RII (TNF R2). These receptors are type I transmembrane glycoproteins belonging to the TNF receptor superfamily, which is defined by the presence of repeating units of cysteine clusters. TNFRI is present on virtually all cell types except for red blood cells. TNFRI is a 55 kDa glycoprotein and contains 5 N-linked glycosylation sites. This is a HCX protein. HCX Expression System Details HCX proteins mimic the proteins in the human body because they are expressed from human, rather than animal, insect or bacterial cells. This process gives them human post-translational modifications. Recombinant DNA techniques allow a human protein with the correct amino acid sequence to be expressed in a non-human cell line. However, non-human cells lack the appropriate cellular machinery, such as specific glycosyltransferases, necessary to produce the correct human post-translational modifications of a protein. An extreme example is seen in E. coli cells, which produce recombinant proteins with no glycosylation, as the above figure illustrates. Rodent and yeast cells are able to glycosylate proteins, but they are still different from glycosylation in human cells. Expression System Resultant Proteins Human (e.g. K562, HEK293) Correct amino acid sequence Human post-translational modifications Rodent (e.g. CHO, NSO) Correct amino acid sequence Some natural glycosylation - not human-like Yeast (e.g. Pichia) Correct amino acid sequence Some natural glycosylation - not human-like E.Coli Correct amino acid sequence No PTMs Although there have been significant attempts to make non-human cell derived cytokines more human-like, there is a growing awareness that in many instances, particularly in therapeutics, cytokines should mimic those found in the body as closely as possible.
Purified TNF RI-Fc HCX Chimera consists of 5-20% carbohydrate by weight.
Limitations
This product is for research use only and is not approved for use in humans or in clinical diagnosis. Peptides and proteins are
guaranteed for 2 years from date of receipt.
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