Product Name: Chk1-pS280
Product Number: AB-PK577
Size: 25 µg      Price:89.00
      $US
Target Full Name: Checkpoint protein-serine kinase 1

Target Alias: CHEK1; CCDS8459.1; ENSG00000149554

Product Type Specific: Protein kinase phosphosite-specific antibody

Antibody Code: PK577

Antibody Target Type: Phosphosite-specific

Antibody Phosphosite: S280

Protein UniProt: O14757

Protein SigNET: O14757

Antibody Type: Polyclonal

Antibody Host Species: Rabbit

Antibody Immunogen Source: Human Chk1 (CHEK1) sequence peptide Cat. No.: PE-04AJV95
Antibody Immunogen Sequence: RVT(pS)GGV(bA)C

Antibody Immunogen Description: Corresponds to amino acid residues R277 to V283; After the kinase catalytic domain in the second half of the kinase

Production Method: The immunizing peptide was produced by solid phase synthesis on a multipep peptide synthesizer and purified by reverse-phase hplc chromatography. Purity was assessed by analytical hplc and the amino acid sequence confirmed by mass spectrometry analysis. This peptide was coupled to KLH prior to immunization into rabbits. New Zealand White rabbits were subcutaneously injected with KLH-coupled immunizing peptide every 4 weeks for 4 months. The sera from these animals was applied onto an agarose column to which the immunogen peptide was thio-linked. Antibody was eluted from the column with 0.1 M glycine, pH 2.5. Subsequently, the antibody solution was neutralized to pH 7.0 with saturated Tris.This antibody was also subject to negative purification over phosphotyrosine-agarose.

Antibody Modification: Unconjugated. Contact KInexus if you are interest in having the antibody biotinylated or coupled with fluorescent dyes.

Antibody Concentration: 1 mg/ml

Storage Buffer: Phosphate buffered saline pH 7.4, 0.05% Thimerasol

Storage Conditions: For long term storage, keep frozen at -40°C or lower. Stock solution can be kept at +4°C for more than 3 months. Avoid repeated freeze-thaw cycles.

Product Use: Western blotting | Antibody microarray

Antibody Dilution Recommended: 2 µg/ml for immunoblotting

Antibody Potency: Weak immunoreactivity of a target-sized protein by Western blotting in HeLa cells with nocodazole overnight treatment. Very strong immunoreactivity with immunogen peptide on dot blots.

Antibody Species Reactivity: Human

Antibody Positive Control: The observed molecular mass of the processed target protein on SDS-PAGE gels is reported to be around 54-58 kDa.

Antibody Specificity: Medium-High

Antibody Cross Reactivity: Nocodazole treatment of HeLa yielded strong increases in 100 as well as 60, 70 and 130 kDa cross-reactive proteins. It is possible that the strong 60 kDa protein is actually the target.

Related Product 1: Chk1-pS280 blocking peptide

Related Product 2: Chk1-pS317 phosphosite-specific antibody (Cat. No.: AB-PK578)

Related Product 3: Chk1-pS345 phosphosite-specific antibody (Cat. No.: AB-PK579)

Related Product 5: Chk1Selectide - Chk1 (CHEK1) protein kinase substrate peptide

Scientific Background: Chk1 (CHEK1) is a protein-serine/threonine kinase of the CAMK group and CAMKL family. It is required for checkpoint-mediated cell cycle arrest. It appears to be a tumour suppressor protein (TSP). The active form of the protein kinase normally acts to inhibit tumour cell proliferation. It activates DNA repair (i.e. homologous recombination) by phosphorylating RAD51 or FANCE in response to the presence of damage or unreplicated DNA. It regulates cell cycle progression via several mechanisms: 1) phosphorylation of certain proteins such as CDC25A, CDC25B and CDC25C, promoting their degradation; 2) phosphorylation and activation of p53; 3) phosphorylation and inactivation of TLK1, which consequently prevents TLK1-dependent phosphorylation of chromatin assembly factor ASF1A; 4) phosphorylation of histones and regulating transcription of genes involved in cell-cycle progression regulation; 5) phosphorylation of RB1 to promote its interaction with E2F transcription factors and subsequent cell cycle arrest. It is nhibited by phosphorylation at S280. S345A mutation of Chk1 leads to reduced phosphorylation and impaired activation to nuclear retention after checkpoint activation. Mutations at 372, 376, and 379 amino acids lead to misfolding and induced binding with ubiquitin.
Figure 1. Dot blotting Chk1-pS280 antibody with recombinant purified proteins.


Figure 2. Epitope mapping of Chk1-pS280 antibody with similar phosphopeptides on dot blots.


Figure 3. Identification of phosphosites related to Chk1-pS280.