The Role of D and E Helices in HCN Channels

Hyperpolarization-activated cyclic-nucleotide regulated (HCN) channels are non-selective cation controlling key physiological functions such as cardiac pacemaking and repetitive neuronal firing. They activated by voltage modulated binding of cAMP to their cytoplasmic C-terminal region which comprises a cyclic nucleotide domain (CNBD). Altered responses caused either point mutations in the protein or non-physiological levels cell lead pathological conditions both heart nervous system. The recently obtained cryo-EM structures human HCN1 cAMP-bound unbound forms confirm previous NMR studies on cAMP-induced conformational changes reveals presence two additional helices at C-terminus CNBD fold cAMP. Inspection structure several putative electrostatic hydrophobic interactions between D E CNBD's C-helix, enable hairpin-like conformation that holds C-helix place. We have previously shown, isothermal titration calorimetry (ITC) patch-clamp experiments, deletion decreases affinity for HCN1, HCN2 HCN4, three HCN isoforms respond Here, we introduce single designed disrupt individual residue with aim investigate molecular mechanism allows fine regulation response. All experiments were performed whole configuration using new all-in-one integrated amplifier, ePatch.