Hochscherf, Jennifer ORCID: 0000-0002-4412-7391, Lindenblatt, Dirk, Steinkrtiger, Michaela, Yoo, Eungyoung, Ulucan, Oezlem, Herzig, Stefan, Issinger, Olaf-Georg, Helms, Volkhard ORCID: 0000-0002-2180-9154, Goetz, Claudia, Neundorf, Ines ORCID: 0000-0001-6450-3991, Niefind, Karsten ORCID: 0000-0002-0183-6315 and Pietsch, Markus (2015). Development of a high-throughput screening-compatible assay to identify inhibitors of the CK2 alpha/CK2 beta interaction. Anal. Biochem., 468. S. 4 - 15. SAN DIEGO: ACADEMIC PRESS INC ELSEVIER SCIENCE. ISSN 1096-0309

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Abstract

Increased activity of protein kinase CK2 is associated with various types of cancer, neurodegenerative diseases, and chronic inflammation. In the search for CK2 inhibitors, attention has expanded toward compounds disturbing the interaction between CK2 alpha and CK2 beta in addition to established active site-directed approaches. The current article describes the development of a fluorescence anisotropy-based assay that mimics the principle of CK2 subunit interaction by using CK2 alpha(1-335) and the fluorescent probe CF-Ahx-Pc as a CK2 beta analog. In addition, we identified new inhibitors able to displace the fluorescent probe from the subunit interface on CK2 alpha(1-335). Both CF-Ahx-Pc and the inhibitors I-Pc and Cl-Pc were derived from the cyclic peptide Pc, a mimetic of the C-terminal CK2 alpha-binding motif of CK2 beta. The design of the two inhibitors was based on docking studies using the known crystal structure of the Pc/CK2 alpha(1-335) complex. The dissociation constants obtained in the fluorescence anisotropy assay for binding of all compounds to human CK2 alpha(1-335) were validated by isothermal titration calorimetry. I-Pc was identified as the tightest binding ligand with a K-D value of 240 nM and was shown to inhibit the CK2 holoenzyme-dependent phosphorylation of PDX-1, a substrate requiring the presence of CK2 beta, with an IC50 value of 92 mu M. (C) 2014 Elsevier Inc. All rights reserved.

Item Type: Journal Article
Creators:
CreatorsEmailORCIDORCID Put Code
Hochscherf, JenniferUNSPECIFIEDorcid.org/0000-0002-4412-7391UNSPECIFIED
Lindenblatt, DirkUNSPECIFIEDUNSPECIFIEDUNSPECIFIED
Steinkrtiger, MichaelaUNSPECIFIEDUNSPECIFIEDUNSPECIFIED
Yoo, EungyoungUNSPECIFIEDUNSPECIFIEDUNSPECIFIED
Ulucan, OezlemUNSPECIFIEDUNSPECIFIEDUNSPECIFIED
Herzig, StefanUNSPECIFIEDUNSPECIFIEDUNSPECIFIED
Issinger, Olaf-GeorgUNSPECIFIEDUNSPECIFIEDUNSPECIFIED
Helms, VolkhardUNSPECIFIEDorcid.org/0000-0002-2180-9154UNSPECIFIED
Goetz, ClaudiaUNSPECIFIEDUNSPECIFIEDUNSPECIFIED
Neundorf, InesUNSPECIFIEDorcid.org/0000-0001-6450-3991UNSPECIFIED
Niefind, KarstenUNSPECIFIEDorcid.org/0000-0002-0183-6315UNSPECIFIED
Pietsch, MarkusUNSPECIFIEDUNSPECIFIEDUNSPECIFIED
URN: urn:nbn:de:hbz:38-419816
DOI: 10.1016/j.ab.2014.09.003
Journal or Publication Title: Anal. Biochem.
Volume: 468
Page Range: S. 4 - 15
Date: 2015
Publisher: ACADEMIC PRESS INC ELSEVIER SCIENCE
Place of Publication: SAN DIEGO
ISSN: 1096-0309
Language: English
Faculty: Unspecified
Divisions: Unspecified
Subjects: no entry
Uncontrolled Keywords:
KeywordsLanguage
PROTEIN-KINASE CK2; CASEIN KINASE-2; BETA-SUBUNIT; FLUORESCENCE POLARIZATION; CRYSTAL-STRUCTURE; 2-PHOTON FLUORESCENCE; PROAPOPTOTIC PEPTIDE; BOMBESIN ANALOGS; BINDING ASSAY; LUNG-CANCERMultiple languages
Biochemical Research Methods; Biochemistry & Molecular Biology; Chemistry, AnalyticalMultiple languages
URI: http://kups.ub.uni-koeln.de/id/eprint/41981

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