PCNA Human, Sf9

Proliferating Cell Antigen Human Recombinant, Sf9
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Cat. No.
BT3024
Source
Sf9 insect cells.
Synonyms
Proliferating cell nuclear antigen, PCNA, Cyclin, MGC8367.
Appearance
Sterile Filtered colorless solution.
Purity
Greater than 95% as determined by SDS-PAGE.
Usage
Prospec's products are furnished for LABORATORY RESEARCH USE ONLY. The product may not be used as drugs, agricultural or pesticidal products, food additives or household chemicals.
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Description

Definition and Production

PCNA Human, Sf9 is a 35 kDa polypeptide chain expressed with a hexahistidine (6xHis) tag in Sf9 cells and purified via proprietary chromatographic techniques . Its catalog number is PRO-303 (Prospec Bio), and it is supplied in a sterile solution containing 16 mM HEPES (pH 8), 400 mM NaCl, and 20% glycerol .

Functional Roles

PCNA acts as a scaffold protein, coordinating DNA replication and repair processes:

  • DNA Replication: Loaded onto DNA by replication factor C (RFC), PCNA tethers DNA polymerase δ (Pol δ) to DNA, enabling processive synthesis of Okazaki fragments .

  • DNA Repair: Facilitates base excision repair and mismatch repair by recruiting repair enzymes .

  • Chromatin Remodeling: Stabilizes interactions between chromatin-modifying enzymes and DNA .

Mutations in PCNA residues (e.g., D41A, R210A) impair RFC binding, ATPase stimulation, and DNA synthesis efficiency .

Experimental Uses

  • Western Blot: Detected using anti-hexa-His-tag antibodies or scleroderma patient sera .

  • ELISA: Coating concentration of 0.5–1.1 µg/ml for autoimmune antibody detection .

  • Structural Studies: Cryo-EM analyses reveal PCNA’s interaction with Pol δ and flap endonuclease 1 (FEN1) during Okazaki fragment processing .

Virology Insights

  • Sf9-derived PCNA colocalizes with baculovirus DNA replication sites, compensating for viral PCNA deficiencies .

Mutational Analyses

MutationFunctional ImpactSource
D41AReduces RFC binding, ATPase activity, and DNA synthesis efficiency by >80%
R210APartially impairs RFC-dependent DNA synthesis but retains RFC-independent activity

Production and Stability

  • Purity: >95% (SDS-PAGE) .

  • Storage: Stable at 4°C for 2–4 weeks or -20°C long-term .

  • Safety: For research use only; not approved for clinical applications .

Product Specs

Introduction
Proliferating cell nuclear antigen (PCNA), a 30 kDa protein also called cyclin, forms a trimer ring encircling a DNA double helix. PCNA interacts with various nuclear proteins, facilitating the organization of biochemical processes at the DNA replication fork. This involvement grants PCNA crucial roles in DNA synthesis, repair, processing, cell cycle regulation, and even apoptosis. Anti-PCNA antibodies are found in systemic lupus erythematosus (SLE), representing a rare specificity (< 10% of SLE patients). Their presence, alone or with other autoantibodies, should alert clinicians to possible existing or developing SLE.
Description
Recombinant Human PCNA, produced in SF9 cells, is a glycosylated polypeptide chain with a molecular weight of 35 kDa. It is expressed with a -6xHis tag and purified using proprietary chromatographic methods.
Physical Appearance
The product appears as a clear, colorless solution that has been sterilized by filtration.
Formulation
Recombinant Human PCNA is provided in a buffer solution consisting of 16mM HEPES (pH 8), 400mM NaCl, and 20% glycerol.
Immunological Functions
This product serves the following immunological functions: (1) It binds to human auto-antibodies of the IgG type. (2) It can be used as a standard in ELISA tests, including checkerboard analysis of positive/negative sera panels and immuno-dot analysis with validated SLE patient sera.
Applications
This product is suitable for use in Western Blot applications with a monoclonal anti-hexa-His-tag antibody and Scleroderma patient sera.
Coating Concentration
The recommended coating concentration for this product is 0.5-1.1 µg/ml. This can vary depending on the ELISA plate type and the coating buffer used. The product is suitable for biotinylation and iodination.
Purity
The purity of this product is greater than 95% as determined by SDS-PAGE analysis.
Stability
For optimal storage, keep at 4°C if the entire vial will be used within 2-4 weeks. For long-term storage, freeze at -20°C. Minimize repeated freeze-thaw cycles.
Synonyms
Proliferating cell nuclear antigen, PCNA, Cyclin, MGC8367.
Source
Sf9 insect cells.

Product Science Overview

Gene and Protein Structure

The PCNA gene is located on human chromosome 20p12.3 and encodes a protein with a molecular weight of approximately 29.8 kDa . The protein is expressed in the nucleus and is involved in various cellular processes, including DNA synthesis, repair, and cell cycle regulation .

Function and Mechanism

PCNA acts as a sliding clamp, providing a stable platform for DNA polymerases and other proteins involved in DNA replication and repair . It binds to a variety of nuclear proteins, organizing biochemical processes at the DNA replication fork . During the cell cycle, PCNA expression peaks during the S-phase, when DNA synthesis occurs, and declines during the G2 and M phases .

Recombinant PCNA in Sf9 Cells

Recombinant PCNA is often produced using the baculovirus expression system in Sf9 insect cells. This system allows for high-level expression of the protein with proper folding and post-translational modifications . The recombinant PCNA is typically tagged with a 6x His tag to facilitate purification using chromatographic techniques .

Applications

Recombinant PCNA is widely used in research to study DNA replication and repair mechanisms. It is also used in assays to identify potential inhibitors of PCNA, which could be valuable in cancer therapy . The protein’s role in maintaining genome stability makes it a critical target for understanding and potentially treating various diseases, including cancer .

Storage and Handling

Recombinant PCNA is usually supplied in a liquid form and should be stored at -70°C to maintain its stability . It is important to avoid freeze-thaw cycles to prevent protein degradation .

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