Water and chloride as allosteric inhibitors in WNK kinase osmosensing

• DOI: 10.7554/elife.88224.1
• Publication Date: 2023-10-23T20:52:11Z
• Authors: Liliana R. Teixeira; Radha Akella; John M. Humphreys; Haixia He; Elizabeth J. Gold...
• Keywords: WNK3; QH301-705.5; Science; Structural Biology and Molecular Biophysics; water; Q; R; Medicine; Biology (General); osmosensor; WNK1; crystallography; chloride ion

Previous observations showed that chloride and osmotic stress regulate the autophosphorylation activity of kinase domains WNK1 WNK3. Further, prior crystallography on asymmetric dimeric unphosphorylated domain (WNK1/S382A, WNK1/SA) revealed conserved waters in active site. Here we show by PEG400 applied to crystals WNK1/SA grown space group P1 induces de-dimerization with a change P2 1 . Both waters, referred here as water network (CWN1) binding site are disrupted PEG400. CWN1 is surrounded stabilized pan-WNK-conserved cluster charged residues. mutagenized these charges WNK3 probe importance WNK regulation. Two mutations at E314 Activation Loop (WNK3/E314Q WNK3/E314A) enhanced activity, consistent idea inhibitory. Mutations other residues had similar or less than wild-type. activation was not significantly reduced point mutants tested. The crystallographic assay data support role for stabilizing an inactive configuration WNKs suggest functions allosteric inhibitor WNKs.