科研之友
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摘要
A new assay for protein kinase CK2 activity determination based on the quantification of a phosphorylated substrate was developed. The common CK2 substrate peptide RRRDDDSDDD, conjugated with the fluorophore 5-[(2-aminoethyl)amino]naphthalene-1-sulfonic acid at the C-terminus served as the analyte. By means of CZE using 2 mol/L acetic acid as electrolyte and UV detection at 214 nm, the non-phosphorylated and the phosphorylated peptide variants could be resolved within 6 min from a complex assay mixture. By this means, activity of human CK2 could be monitored by a kinetic, as well as an endpoint, method. inhibition of human recombinant CK2 holoenzyme by 6-methyl-1, 3,8-trihydroxyanthraquinone and 4,5,6,7-tetrabromobenzotriazole resulted in IC(50) values of 1.33 and 0.27 mu M, respectively, which were similar to those obtained with the standard radiometric assay. These results suggest that the CE/UV strategy described here is a straightforward assay for CK2 inhibitor testing.
- 出版日期2010-2
