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...
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MDPI AG
2017-01-01
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| Series: | Pharmaceuticals |
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| Online Access: | http://www.mdpi.com/1424-8247/10/1/6 |
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| author | Christian Nienberg Claudia Garmann Andreas Gratz Andre Bollacke Claudia Götz Joachim Jose |
| author_facet | Christian Nienberg Claudia Garmann Andreas Gratz Andre Bollacke Claudia Götz Joachim Jose |
| author_sort | Christian Nienberg |
| collection | DOAJ |
| description | 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 × 105 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 CK2α subunit (IC50 = 0.8 µM). 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 CK2α was determined to be 2.16 µM, whereas no binding of B2 to CK2β-subunit was detectable. To our surprise, besides inhibition of enzymatic activity, B2 also disturbed the interaction of CK2α with CK2β at higher concentrations (≥25 µM). |
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| institution | Directory Open Access Journal |
| issn | 1424-8247 |
| language | English |
| last_indexed | 2024-12-23T11:28:57Z |
| publishDate | 2017-01-01 |
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| spelling | doaj.art-b394dc6f196b47afa0d2e04c9dd1395a2022-12-21T17:48:51ZengMDPI AGPharmaceuticals1424-82472017-01-01101610.3390/ph10010006ph10010006Identification of a Potent Allosteric Inhibitor of Human Protein Kinase CK2 by Bacterial Surface Display Library ScreeningChristian Nienberg0Claudia Garmann1Andreas Gratz2Andre Bollacke3Claudia Götz4Joachim Jose5Institut für Pharmazeutische und Medizinische Chemie, PharmaCampus, Westfälische Wilhelms-Universität Münster, Correnstraße 48, D-48149 Münster, GermanyInstitut für Pharmazeutische und Medizinische Chemie, PharmaCampus, Westfälische Wilhelms-Universität Münster, Correnstraße 48, D-48149 Münster, GermanyInstitut für Pharmazeutische und Medizinische Chemie, PharmaCampus, Westfälische Wilhelms-Universität Münster, Correnstraße 48, D-48149 Münster, GermanyInstitut für Pharmazeutische und Medizinische Chemie, PharmaCampus, Westfälische Wilhelms-Universität Münster, Correnstraße 48, D-48149 Münster, GermanyMedizinische Biochemie und Molekularbiologie, Universität des Saarlandes, Kirrberger Str., Geb. 44, D-66421 Homburg, GermanyInstitut für Pharmazeutische und Medizinische Chemie, PharmaCampus, Westfälische Wilhelms-Universität Münster, Correnstraße 48, D-48149 Münster, GermanyHuman 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 × 105 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 CK2α subunit (IC50 = 0.8 µM). 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 CK2α was determined to be 2.16 µM, whereas no binding of B2 to CK2β-subunit was detectable. To our surprise, besides inhibition of enzymatic activity, B2 also disturbed the interaction of CK2α with CK2β at higher concentrations (≥25 µM).http://www.mdpi.com/1424-8247/10/1/6autodisplayhuman protein kinase CK2non ATP-competitive inhibitorpeptide |
| spellingShingle | Christian Nienberg Claudia Garmann Andreas Gratz Andre Bollacke Claudia Götz Joachim Jose Identification of a Potent Allosteric Inhibitor of Human Protein Kinase CK2 by Bacterial Surface Display Library Screening Pharmaceuticals autodisplay human protein kinase CK2 non ATP-competitive inhibitor peptide |
| title | Identification of a Potent Allosteric Inhibitor of Human Protein Kinase CK2 by Bacterial Surface Display Library Screening |
| title_full | Identification of a Potent Allosteric Inhibitor of Human Protein Kinase CK2 by Bacterial Surface Display Library Screening |
| title_fullStr | Identification of a Potent Allosteric Inhibitor of Human Protein Kinase CK2 by Bacterial Surface Display Library Screening |
| title_full_unstemmed | Identification of a Potent Allosteric Inhibitor of Human Protein Kinase CK2 by Bacterial Surface Display Library Screening |
| title_short | Identification of a Potent Allosteric Inhibitor of Human Protein Kinase CK2 by Bacterial Surface Display Library Screening |
| title_sort | identification of a potent allosteric inhibitor of human protein kinase ck2 by bacterial surface display library screening |
| topic | autodisplay human protein kinase CK2 non ATP-competitive inhibitor peptide |
| url | http://www.mdpi.com/1424-8247/10/1/6 |
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