Identification of a Potent Allosteric Inhibitor of Human Protein Kinase CK2 by Bacterial Surface Display Library Screening.

Human protein kinase CK2 has emerged as promising target for the treatment of neoplastic diseases. The vast majority of kinase inhibitors known today target the ATP binding site, which is highly conserved among kinases and hence leads to limited selectivity. In order to identify non-ATP competitive inhibitors, a 12-mer peptide library of 6 x 10(5) variants was displayed on the surface of E. coli by autodisplay. Screening of this peptide library on variants with affinity to CK2 was performed by fluorophore-conjugated CK2 and subsequent flow cytometry. Single cell sorting of CK2-bound E. coli yielded new peptide variants, which were tested on inhibition of CK2 by a CE-based assay. Peptide B2 (DCRGLIVMIKLH) was the most potent inhibitor of both, CK2 holoenzyme and the catalytic CK2 alpha subunit (IC50 = 0.8 mu M). Using different ATP concentrations and different substrate concentrations for IC50 determination, B2 was shown to be neither ATP-nor substrate competitive. By microscale thermophoresis (MST) the KD value of B2 with CK2 alpha was determined to be 2.16 mu M, whereas no binding of B2 to CK2 beta-subunit was detectable. To our surprise, besides inhibition of enzymatic activity, B2 also disturbed the interaction of CK2 alpha with CK2 beta at higher concentrations (>= 25 mu M).