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Updated: Jul 29, 2026
08:05
Assaying Protein Kinase Activity with Radiolabeled ATP
Published on: May 26, 2017
Characterization of PDZ-binding kinase, a mitotic kinase.
1Department of Molecular and Cellular Biology, Harvard University, 16 Divinity Avenue, Cambridge, MA 02138, USA.
Summary
A novel serine/threonine kinase, PDZ-binding kinase (PBK), interacts with the hDlg tumor suppressor protein. PBK
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- The human homologue of Drosophila Discs-large (hDlg) is a tumor suppressor protein known to interact with APC and HPV E6.
- PDZ domains are crucial protein interaction modules involved in scaffolding and signal transduction.
Purpose of the Study:
- To identify novel binding partners of the hDlg protein.
- To characterize the function and regulation of a newly identified kinase interacting with hDlg.
Main Methods:
- Yeast two-hybrid screening to identify hDlg-interacting proteins.
- In vitro binding assays using purified proteins.
- Western blotting and kinase assays in HeLa cells.
- In vitro phosphorylation assays using cdc2/cyclin B.
Main Results:
- A novel serine/threonine kinase, PDZ-binding kinase (PBK), was identified that binds to the PDZ2 domain of hDlg via a C-terminal T/SXV motif.
- PBK mRNA is highly expressed in placenta and absent in adult brain.
- Both PBK and hDlg are phosphorylated during mitosis.
- Mitotic phosphorylation of PBK is essential for its kinase activity, and cdc2/cyclin B phosphorylates PBK in vitro.
Conclusions:
- PBK is a novel kinase that binds to hDlg, suggesting a role in regulating cell cycle or proliferation.
- Mitotic phosphorylation by cdc2/cyclin B regulates PBK activity.
- PBK may link hDlg and other PDZ-containing proteins to cell cycle regulatory pathways.
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