Insight into the Selectivity of Kir3.2 toward Phosphatidylinositides

• DOI: 10.1021/acs.biochem.0c00163
• Publication Date: 2020-05-06T10:47:28Z
• Authors: Pei Qiao; Yang Liu; Tianqi Zhang; Amanda Benavides; Arthur Laganowsky
• Keywords: 0301 basic medicine; Mice; 03 medical and health sciences; G Protein-Coupled Inwardly-Rectifying Potassium Channels; Fluorescence Resonance Energy Transfer; Animals; Point Mutation; Phosphatidylinositols; Mass Spectrometry

Activation of G-protein-gated inwardly rectifying potassium channels (Kir3.x) requires the direct binding phosphorylated phosphatidylinositides (PIPs). Previous studies have established that PIP isoforms activate Kir to varying degrees and affinity between PIPs Kir3.2 appears be correlated with level activation. However, how individual residues contribute selectivity toward is poorly understood. Here, we employ native mass spectrometry (MS) fluorescent lipid assays gain insight into contribution specific phospholipids. For wild-type channel, demonstrate importance membrane protein samples devoid co-purified contaminants for protein–lipid show PIP(4,5)P2 cooperatively binds a Hill coefficient 2.7. We also find profiles determined from MS solution are in agreement. Point mutations interact distinctly alter selective binding. The K64Q mutation results altered highest acyl chains. Mutation R92 Pro, residue found Kir6.2, promiscuous isoforms. K194A distinct preference PIP(3,4,5)P3 over other Taken together, our underscore utmost quality single can PIPs.