Furin Overexpression Lysate

Novus Biologicals | Catalog # NBL1-10860

Novus Biologicals
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Key Product Details

Species

Human

Applications

Western Blot

Product Summary for Furin Overexpression Lysate

Furin Transient Overexpression Lysate


Expression Host: HEK293T

Plasmid: RC204279

Accession#: NM_002569

Protein Tag: C-MYC/DDK

You will receive 1 vial of lysate (100ug), 1 vial of empty vector negative control (100ug), and 1 vial of 2xSDS sample buffer (250ul). Each vial of cell lysate contains 100ug of total protein (at 1 mg/ml). The 2xSDS Sample Buffer consists of 4% SDS, 125mM Tris-HCl pH6.8, 10% Glycerol, 0.002% Bromophenol blue, 100mM DTT.

Product Specifications

Application Notes

This product is intended for use as a positive control in Western Blot. Overexpression of the target protein was confirmed using an antibody to DDK (FLAG) epitope tag (NBP1-71705) present on the protein construct.

Each vial of cell lysate contains 100ug of total protein which should be sufficient for 20-50 reactions. Depending on over-expression level, antibody affinity and detection system, some lysates can go as low as 0.1 ug per load. We recommend starting with 5ug of cell lysate. Add an equal amount of cell lysate and 2X SDS Sample buffer and boil the SDS samples for 10 minutes before loading.

Theoretical MW

86.5 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Type

Overexpression

Scientific Data Images for Furin Overexpression Lysate

Western Blot: Furin Overexpression Lysate [NBL1-10860]

Western Blot: Furin Overexpression Lysate [NBL1-10860]

Western Blot: Furin Overexpression Lysate (Adult Normal) [NBL1-10860] Left-Empty vector transfected control cell lysate (HEK293 cell lysate); Right -Over-expression Lysate for Furin.

Formulation, Preparation, and Storage

Formulation

RIPA buffer

Concentration

The exact concentration of the protein of interest cannot be determined for overexpression lysates. Please contact technical support for more information.

Shipping

The product is shipped with dry ice or equivalent. Upon receipt, store it immediately at the temperature recommended below.

Storage

Store at -80C. Avoid freeze-thaw cycles.

Background: Furin

Furin (also known as FUR, PACE, PCSK3, and SCP1) is a ubiquity expressed, calcium-dependent serine protease that belongs to the subtilisin-like proprotein convertase (PC) family (1). Furin is synthesized as a 794-amino acid type-I transmembrane protein (1). Like other PC family members, furin shares structural similarity which includes a heterogeneous ~10 kDa amino terminal proregion, a ~55 kDa highly conserved subtilisin-like catalytic domain, and a carboxylterminal domain that is heterogeneous and varies in length and sequence between different PC family members (2). Furin plays an important role in embryogenesis and assists in the maturation of proprotein substrates (1,3). As a protease, furin cleaves and activates over 150 mammalian, viral, and bacterial substrates (3). These substrates include growth factors, receptors, hormones, cytokines, and adhesion molecules (1, 3). Furin processes these proproteins in secretory pathway compartments by cleaving at the carboxyl-terminus of the consensus sequence RX(K/R)R (where X is any amino acid) (1). Furin is localized to the trans-Golgi-network by the acidic peptide sequence C771PSDSEEDEG780 where phosphorylation of the serine residues regulates the intracellular trafficking. Another unique signaling domain is the hydrophobic motif Y759KGL762 that modulates endocytosis from the cell surface. The theoretical molecular weight of furin is 87 kDa; however, the observed molecular weight may vary as post-translational modifications, like glycosylation, cause the pre-pro furin to run at 110-104 kDa, the mature furin at 98-95 kDa, and the shed furin at 90 kDa (4).

Proteolytic cleavage regulates several physiological processes in both health and disease (3). Abnormal activity or mutations in proteases, including furin, is associated with pathologies and diseases including cancer, cardiovascular disorders, diabetes, inflammation, neurological diseases, and autoimmune diseases (3). As mentioned above, furin also acts upon bacterial substrates, including anthrax and Shiga toxin, and many virus families such as Herpes-, Flavi-, and Corona-, leading to host infections. Furthermore, the novel coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) present with a S-spike protein that is cleaved by PCs, including furin, at the S1/S2 cleavage site (5, 6). The cleavage allows the SARS-CoV-2 to then attach to the angiotensin-converting enzyme 2 (ACE2) receptor via the S1 domain and the cellular membrane via the S2 domain (5, 6). Although COVID-19 patients mostly present with respiratory symptoms, a variety of other systems are affected including cardiovascular, gastrointestinal (GI), and the liver (5-7). It is suggested that the S1/furin/ACE2 interaction promotes SARS-CoV-2 infection leading to the harmful symptoms and reactions in patients (5, 6). Cardiovascular disease is a common comorbidity in patients, along with hypertension, myocardial damage, and heart palpitations (Ming). Further evidence of furin being a risk factor for infection is the high levels of furin present in the blood of heart failure patients (5). Similarly, the small bowel may be another interaction site for infection as it is rich in furin and the intestinal enterocytes have many ACE2 receptors (6). Furin is also highly expressed in the liver and hepatocytes and cholangiocytes of the liver present ACE2 receptors (3, 7). Studies have shown that up one-third of COVID-19 patients experience GI symptoms which range from diarrhea and loss of appetite to abdominal cramping and bloody stool (6, 7). Additionally, some patients displayed abnormal liver enzyme levels (7). It has been suggested that a possible therapeutic strategy for treating those infected with SARS-CoV-2 is pharmacologically or immunologically modulating furin or ACE2 binding sites to combat COVID-19 infection (3, 5).

References

1. Thomas, G. (2002). Furin at the cutting edge: from protein traffic to emryogenesis and disease. Nature Rev. Mol. Cell Biol. https://doi.org/10.1038/nrm934

2. Zhou A., Paquet, L., & Mains, R.E. (1995). Structural elements that direct specific processing of different mammalian subtilisin-like prohormone convertases. J Biol Chem. https://doi.org/10.1074/jbc.270.37.21509

3. Braun E., & Sauter, D. (2019). Furin-mediated protein processing in infectious diseases and cancer. Clin Transl Immunology. https://doi:10.1002/cti2.1073

4. Atlas of Genetics and Cytogenetics in Oncology and Haematology, FURIN

5. Ming, Y. & Qiang, L. (2020). Involvement of Spike Protein, Furin, and ACE2 in SARS-CoV-2-Related Cardiovascular Complications. SN Compr. Clin. Med. https://doi.org/10.1007/s42399-020-00400-2

6. Monkemuller, K., Fry, L., & Rickes, S. (2020). COVID-19, coronavirus, SARS-CoV-2 and the small bowel. Rev Esp Enferm Dig. https://doi:10.17235/reed.2020.7137/2020

7. Agarwal, A., Chen, A., Ravindran, N., To, C., & Thuluvath, P.J. (2020). Gastrointestinal and Liver Manifestations of COVID-19. J Clin Exp Hepatol. https://doi:10.1016/j.jceh.2020.03.001

Alternate Names

FUR, PACE, PCSK3, SPC1

Gene Symbol

FURIN

Additional Furin Products

Product Documents for Furin Overexpression Lysate

Certificate of Analysis

To download a Certificate of Analysis, please enter a lot or batch number in the search box below.

Product Specific Notices for Furin Overexpression Lysate

HEK293T cells in 10-cm dishes were transiently transfected with a non-lipid polymer transfection reagent specially designed and manufactured for large volume DNA transfection. Transfected cells were cultured for 48hrs before collection. The cells were lysed in modified RIPA buffer (25mM Tris-HCl pH7.6, 150mM NaCl, 1% NP-40, 1mM EDTA, 1xProteinase inhibitor cocktail mix, 1mM PMSF and 1mM Na3VO4, and then centrifuged to clarify the lysate. Protein concentration was measured by BCA protein assay kit.

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Lysates are guaranteed for 6 months from date of receipt.

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Associated Pathways

Notch Signaling Pathways Notch Signaling Pathway Thumbnail