Atomic-level investigation of KCNJ2 mutations associated with ventricular arrhythmic syndrome phenotypes.

KCNJ2 encodes the inward rectifying potassium channel (Kir2.1) that underlies I which maintains the cardiac resting membrane potential and regulates excitability. Mutations in KCNJ2 have been linked to several clinical phenotypes associated with life-threatening ventricular arrhythmia and sudden death including Andersen-Tawil syndrome (ATS) from loss of function mutations, and Short QT Syndrome 3 from gain of function mutations. Detailed structural-functional relationships to explain the arrhythmia phenotypes are understudied and limit the capacity to provide precision medicine. Here, we combine in-depth and complementary computational molecular modeling techniques with functional analysis from three patients with ATS that harbor KCNJ2 mutations R67Q, R218L, and G300D. Whole-cell patch-clamp experiments revealed loss of function in homomeric mutant channels. Full-length Kir2.1 models were developed for structure-based investigation, and mutations were introduced in both open and closed conformations. Site-directed mutagenesis identified altered interaction profiles contributing to structural perturbations. Molecular dynamics simulations assessed the impact of each mutation on overall channel conformation and stability. Principal component analysis and normal mode analysis revealed mutation-specific structural perturbations. The findings afford atomic mechanistic underpinnings of mutation-specific perturbations. Our multifaceted approach provides first atomic-level insights into the molecular mechanisms underlying ATS, paving the way for structure-guided targeted therapeutic strategies for ATS and related channelopathies.