Name: Recombinant Human Active Chk1 Protein, CF
Brand: Novus Biologicals
SKU: 1630-KS
Price: 508 USD

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At	=	A. thaliana	SyHa	=	Golden Syrian Hamster
All	=	All Species	Ha	=	Hamster
An	=	Animal	Hu	=	Human
ArHa	=	Armenian Hamster	I	=	Insect
Av	=	Avian	Ll	=	Llama
Bb	=	Baboon	Ma	=	Mammal
Ba	=	Bacteria	Mk	=	Monkey
Bv	=	Bovine	Mu	=	Mouse
Ca	=	Canine	Multi	=	Multi-species
Ce	=	C. Elegans	NA	=	Non-species specific
Ch	=	Chicken	Or	=	Orangutan
ChHa	=	Chinese Hamster	Pl	=	Plant
Cm	=	Cynomolgus Monkey	Pm	=	Primate
Do	=	Donkey	Po	=	Porcine
Dr	=	Drosophilia	Rb	=	Rabbit
Ec	=	E. Coli	Rt	=	Rat
Eq	=	Equine	RM	=	Rhesus Macaque
Fe	=	Feline	Sh	=	Sheep
Ft	=	Ferret	Vb	=	Vertebrate
Fi	=	Fish	Vi	=	Virus
Fu	=	Fungi	Xp	=	Xenopus
Ge	=	Gerbil	Ye	=	Yeast
GP	=	Guinea Pig	Ze	=	Zebrafish
Gt	=	Goat

Reactivity	HuSpecies Glossary
Applications	Bioactivity
Format	Carrier-Free

Details of Functionality	The specific activity of Chk1 was determined to be 228 nmol/min/mg using a synthetic peptide substrate (KKKVSRSGLYRSPSMPENLNRPR).
Source	Spodoptera frugiperda, Sf 9 (baculovirus)-derived human Chk1 protein
Accession #	NM_001274
N-terminal Sequence	Using an N-terminal GST tag
Protein/Peptide Type	Recombinant Enzymes
Gene	CHEK1
Purity	>95%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane.

Storage	This product is stable at ≤ ‑70°C for up to 1 year from the date of receipt. For optimal storage, aliquot into smaller quantities after centrifugation and store at recommended temperature. Avoid repeated freeze-thaw cycles.
Buffer	Supplied in 50 mM Tris-HCl, pH 7.5, 150 mM NaCl, 10 mM Glutathione, 0.1 mM EDTA, 0.25 mM DTT, 0.1 mM PMSF, 25% glycerol.
Purity	>95%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane.
Assay Procedure	Active Kinase - Active Chk1 (0.1 μg/μL) diluted with Kinase Dilution Buffer I to the concentrations shown in Figure 2. Note: These are suggested working dilutions. Optimal dilutions should be determined by each laboratory for each application. Kinase Assay Buffer I - 25 mM MOPS, pH 7.2, 12.5 mM beta -glycerol-phosphate, 25 mM MgCl₂, 5 mM EGTA, 2 mM EDTA. Add 0.25 mM DTT to the Kinase Assay Buffer I prior to use. Kinase Dilution Buffer - Kinase Assay Buffer I diluted at a 1:4 ratio (5-fold) with distilled water. 10 mM ATP Stock Solution - Prepare the ATP Stock Solution by dissolving 55 mg of ATP in 10 mL of Kinase Assay Buffer I. [³³P]-ATP Assay Cocktail - Prepare 250 μM [³³P]-ATP Assay Cocktail in a designated radioactive work area by combining 150 μL of 10 mM ATP Stock Solution, 100 μL of [³³P]-ATP (1 mCi/100 μL), and 5.75 mL of Kinase Assay Buffer I. Store 1.0 mL aliquots at ≤ -20 °C. Substrate - Chk-tide synthetic peptide substrate (KKKVSRSGLRSPSMPENLNRPR) diluted in distilled or deionized water to a final concentration of 1 mg/mL. Thaw the [³³P]-ATP Assay Cocktail in a shielded container in a designated radioactive work area. Thaw the Active Chk1, Kinase Assay Buffer, Substrate, and Kinase Dilution Buffer on ice. In a pre-cooled microfuge tube, add the following reaction components bringing the initial reaction volume up to 20 μL. a. Diluted Active Chk1: 10 μL b. Substrate (1 mg/mL Stock Solution): 5.0 μL c. Distilled water: 5.0 μL Set up the blank control as outlined in step 3, excluding the addition of the substrate. Replace the substrate with an equal volume of distilled water. Initiate the reaction by the addition of 5 μL [³³P]-ATP Assay Cocktail, bringing the final volume up to 25 μL. Incubate the mixture in a water bath at 30 °C for 15 minutes. After the 15 minute incubation, terminate the reaction by spotting 20 μL of the reaction mixture onto individual pre-cut strips of phosphocellulose P81 paper. Air dry the pre-cut P81 strip and sequentially wash in a 1% phosphoric acid solution (add 10 mL of phosphoric acid to 990 mL of distilled water) with constant gentle stirring. It is recommended that the strips be washed a total of three times for approximately 10 minutes each. Count the radioactivity on the P81 paper in the presence of scintillation fluid in a scintillation counter. Determine the corrected cpm by removing the blank control value (see step 4) for each sample and calculate the kinase specific activity as outlined below. Calculation of [³³P]-ATP Specific Activity (SA) (cpm/pmol) Specific Activity (SA) = cpm for 5 μL [³³P]-ATP/pmol of ATP (in 5 μL of a 250 μM ATP stock solution; i.e.1250 pmol) Calculation of Kinase Specific Activity (SA) (pmol/minutes/μg or nmol/minutes/mg) Corrected cpm from reaction / [(SA of ³³P-ATP in cpm/pmol) x (Reaction time in minutes) x (Enzyme amount in μg or mg)] x [(Reaction volume) / (Spot Volume)]

Publications using 1630-KS	Applications	Species
B Peng, R Shi, J Bian, Y Li, P Wang, H Wang, J Liao, WG Zhu, X Xu PARP1 and CHK1 coordinate PLK1 enzymatic activity during the DNA damage response to promote homologous recombination-mediated repair Nucleic Acids Research, 2021-07-21;49(13):7554-7570. 2021-07-21 [PMID: 34197606] (Bioassay, Human)	Bioassay	Human
R Yuan, HR Vos, RM van Es, J Chen, BM Burgering, B Westendorp, A de Bruin Chk1 and 14-3-3 proteins inhibit atypical E2Fs to prevent a permanent cell cycle arrest EMBO J., 2018-01-23;0(0):. 2018-01-23 [PMID: 29363506] (Bioassay, Human)	Bioassay	Human
Z Li, X Li, S Nai, Q Geng, J Liao, X Xu, J Li Checkpoint kinase 1-induced phosphorylation of O-linked ?-N-acetylglucosamine transferase regulates the intermediate filament network during cytokinesis J. Biol. Chem., 2017-10-11;0(0):. 2017-10-11 [PMID: 29021254] (Bioassay)	Bioassay

Recombinant Human Active Chk1 Protein, CF

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Bioinformatics

Gene Symbol	CHEK1
Uniprot	NM_001274(Human)