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Co-crystal structures of inhibitors with MRCKβ, a key regulator of tumor cell invasion

DOI: 10.1371/journal.pone.0024825 DOI Help
PMID: 21949762 PMID Help

Authors: Timo Heikkila (Wolfson Institute for Biomedical Research) , Edward Wheatley (Wolfson Institute for Biomedical Research) , Diane Crighton (Beatson Institute for Cancer Research) , Ewald Schroder (Wolfson Institute for Biomedical Research) , Alexandra Boakes (Wolfson Institute for Biomedical Research) , Sarah J. Kaye (Wolfson Institute for Biomedical Research) , Mokdad Mezna (Beatson Institute for Cancer Research) , Leon Pang (Wolfson Institute for Biomedical Research) , Mathew Rushbrooke (Wolfson Institute for Biomedical Research) , Andrew Turnbull (Wolfson Institute for Biomedical Research) , Michael F. Olson (Beatson Institute for Cancer Research)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Plos One , VOL 6 (9)

State: Published (Approved)
Published: September 2011

Open Access Open Access

Abstract: MRCKα and MRCKβ (myotonic dystrophy kinase-related Cdc42-binding kinases) belong to a subfamily of Rho GTPase activated serine/threonine kinases within the AGC-family that regulate the actomyosin cytoskeleton. Reflecting their roles in myosin light chain (MLC) phosphorylation, MRCKα and MRCKβ influence cell shape and motility. We report further evidence for MRCKα and MRCKβ contributions to the invasion of cancer cells in 3-dimensional matrix invasion assays. In particular, our results indicate that the combined inhibition of MRCKα and MRCKβ together with inhibition of ROCK kinases results in significantly greater effects on reducing cancer cell invasion than blocking either MRCK or ROCK kinases alone. To probe the kinase ligand pocket, we screened 159 kinase inhibitors in an in vitro MRCKβ kinase assay and found 11 compounds that inhibited enzyme activity >80% at 3 µM. Further analysis of three hits, Y-27632, Fasudil and TPCA-1, revealed low micromolar IC50 values for MRCKα and MRCKβ. We also describe the crystal structure of MRCKβ in complex with inhibitors Fasudil and TPCA-1 bound to the active site of the kinase. These high-resolution structures reveal a highly conserved AGC kinase fold in a typical dimeric arrangement. The kinase domain is in an active conformation with a fully-ordered and correctly positioned αC helix and catalytic residues in a conformation competent for catalysis. Together, these results provide further validation for MRCK involvement in regulation of cancer cell invasion and present a valuable starting point for future structure-based drug discovery efforts.

Journal Keywords: Amides; Catalytic; Cell; Tumor; Collagen; Crystallography; X-Ray; Drug; Extracellular; Humans; Inhibitory; Laminin; Models; Molecular; Myotonin-Protein; Neoplasm; Protein; Protein-Tyrosine; Proteoglycans; Pyridines; Thiophenes; rho-Associated Kinases

Subject Areas: Biology and Bio-materials, Medicine

Instruments: I04-Macromolecular Crystallography

Added On: 25/10/2011 12:09


Discipline Tags:

Non-Communicable Diseases Health & Wellbeing Cancer Structural biology Drug Discovery Life Sciences & Biotech

Technical Tags:

Diffraction Macromolecular Crystallography (MX)