9zkg
VRK1 in complex with the inhibitor MP-60
Structural highlights
DiseaseVRK1_HUMAN Pontocerebellar hypoplasia type 1. The disease is caused by mutations affecting the gene represented in this entry. FunctionVRK1_HUMAN Serine/threonine kinase involved in Golgi disassembly during the cell cycle: following phosphorylation by PLK3 during mitosis, required to induce Golgi fragmentation. Acts by mediating phosphorylation of downstream target protein. Phosphorylates 'Thr-18' of p53/TP53 and may thereby prevent the interaction between p53/TP53 and MDM2. Phosphorylates casein and histone H3. Phosphorylates BANF1: disrupts its ability to bind DNA, reduces its binding to LEM domain-containing proteins and causes its relocalization from the nucleus to the cytoplasm. Phosphorylates ATF2 which activates its transcriptional activity.[1] [2] [3] [4] [5] [6] Publication Abstract from PubMedVaccinia-related kinase 1 (VRK1) is a promising therapeutic target in gliomas and glioblastomas where VRK2 is silenced by promoter methylation, rendering VRK1 essential for accurate nuclear envelope reassembly following mitosis. Small-molecule ATP-site drug discovery for VRK1 has been hindered by the absence of robust and reproducible biochemical assays. Through virtual screening, we identified previously unreported VRK1-binding scaffolds and validated them in biochemical kinase assays, yielding an 82 nM inhibitor with high selectivity for VRK1 over VRK2. During characterization of this compound, we found that a commonly used commercial time-resolved fluorescence resonance energy transfer (TR-FRET) VRK1 activity assay is dependent on purification tag-mediated VRK1 dimerization. Leveraging the new inhibitor, we developed fluorogenic tool compounds that increase in fluorescence intensity upon binding to the active site of VRK1, and do not require artificial dimerization of VRK1. The top probe exhibits a K(d) of 180 nM and is useful for ligand displacement assays using both fluorescence enhancement and TR-FRET readouts. Together, these results introduce new chemical scaffolds for targeting VRK1, define an assay artifact that has complicated VRK1 inhibitor discovery, and deliver fluorogenic tool compounds for high-throughput screening of ATP-site VRK1 inhibitors, enabling future drug discovery efforts against this emerging cancer vulnerability. Structure-Based Discovery of Selective Vaccinia-Related Kinase 1 Inhibitors and Fluorogenic Active-Site Probes.,Crowley-Dolen EK, Borges RJ, Charifson PS, Payne NC, de Oliveira RG, Cunha MR, Mazitschek R, Massirer KB, Mitchison TJ J Biol Chem. 2026 Mar 6:111355. doi: 10.1016/j.jbc.2026.111355. PMID:41796804[7] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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