Nek7 conformational flexibility and inhibitor binding probed through protein engineering of the R-spine
| Autor: | Chitra Bhatia, Katherine H. Carr, Sharon Yeoh, Corine Mas-Droux, Richard Bayliss, Rory F. Cunnison, Matthew J. Byrne, Nazia Nasir, Celine Cano, Christine Basmadjian |
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| Rok vydání: | 2020 |
| Předmět: |
Protein Conformation
Stereochemistry Mutant Substituent Biochemistry Catalysis Serine 03 medical and health sciences chemistry.chemical_compound 0302 clinical medicine Structural Biology Chemical Biology Humans NIMA-Related Kinases Enzyme Inhibitors Molecular Biology Research Articles 030304 developmental biology 0303 health sciences Chemistry Kinase protein engineering Cell Biology Protein engineering Small molecule Kinetics small molecules kinases 030220 oncology & carcinogenesis Mutation Multipolar spindles Cytokinesis Protein Binding |
| Zdroj: | 'Biochemical Journal ', vol: 477, pages: 1525-1539 (2020) Biochemical Journal |
| ISSN: | 1470-8728 0264-6021 |
| Popis: | Nek7 is a serine/threonine-protein kinase required for proper spindle formation and cytokinesis. Elevated Nek7 levels have been observed in several cancers, and inhibition of Nek7 might provide a route to the development of cancer therapeutics. To date, no selective and potent Nek7 inhibitors have been identified. Nek7 crystal structures exhibit an improperly formed regulatory-spine (R-spine), characteristic of an inactive kinase. We reasoned that the preference of Nek7 to crystallise in this inactive conformation might hinder attempts to capture Nek7 in complex with Type I inhibitors. Here, we have introduced aromatic residues into the R-spine of Nek7 with the aim to stabilise the active conformation of the kinase through R-spine stacking. The strong R-spine mutant Nek7SRS retained catalytic activity and was crystallised in complex with compound 51, an ATP-competitive inhibitor of Nek2 and Nek7. Subsequently, we obtained the same crystal form for wild-type Nek7WT in apo form and bound to compound 51. The R-spines of the three well-ordered Nek7WT molecules exhibit variable conformations while the R-spines of the Nek7SRS molecules all have the same, partially stacked configuration. Compound 51 bound to Nek2 and Nek7 in similar modes, but differences in the precise orientation of a substituent highlights features that could be exploited in designing inhibitors that are selective for particular Nek family members. Although the SRS mutations are not required to obtain a Nek7–inhibitor structure, we conclude that it is a useful strategy for restraining the conformation of a kinase in order to promote crystallogenesis. |
| Databáze: | OpenAIRE |
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