@article {Kondapuram2020.09.21.305797, author = {Mahesh Kondapuram and Benedikt Frieg and Sezin Y{\"u}ksel and Tina Schwabe and Christian Sattler and Marco Lelle and Andrea Schweinitz and Ralf Schmauder and Klaus Benndorf and Holger Gohlke and Jana Kusch}, title = {Functional and structural characterization of interactions between opposite subunits in HCN pacemaker channels}, elocation-id = {2020.09.21.305797}, year = {2020}, doi = {10.1101/2020.09.21.305797}, publisher = {Cold Spring Harbor Laboratory}, abstract = {Hyperpolarization-activated and cyclic nucleotide (HCN) modulated channels are tetrameric cation channels. In each of the four subunits, the intracellular cyclic nucleotide-binding domain (CNBD) is coupled to the transmembrane domain via a helical structure, the C-linker. High-resolution channel structures suggest that the C-linker enables functionally relevant interactions with the opposite subunit, which might be critical for coupling the conformational changes in the CNBD to the channel pore. We combined mutagenesis, patch-clamp technique, confocal patch-clamp fluorometry, and molecular dynamics simulations to show that residue K464 of the C-linker is essential for stabilizing the closed state of the mHCN2 channel by forming interactions with the opposite subunit. MD simulations revealed that both cAMP and K464E induce a rotation of the intracellular domain relative to the channel pore, weakening the autoinhibitory effect of the unoccupied CL-CNBD region. The adopted poses are in excellent agreement with structural results.Competing Interest StatementThe authors have declared no competing interest.}, URL = {https://www.biorxiv.org/content/early/2020/09/21/2020.09.21.305797}, eprint = {https://www.biorxiv.org/content/early/2020/09/21/2020.09.21.305797.full.pdf}, journal = {bioRxiv} }