Prolyl isomerization controls activation kinetics of a cyclic nucleotide-gated ion channel

Nat Commun. 2020 Dec 16;11(1):6401. doi: 10.1038/s41467-020-20104-4.


SthK, a cyclic nucleotide-modulated ion channel from Spirochaeta thermophila, activates slowly upon cAMP increase. This is reminiscent of the slow, cAMP-induced activation reported for the hyperpolarization-activated and cyclic nucleotide-gated channel HCN2 in the family of so-called pacemaker channels. Here, we investigate slow cAMP-induced activation in purified SthK channels using stopped-flow assays, mutagenesis, enzymatic catalysis and inhibition assays revealing that the cis/trans conformation of a conserved proline in the cyclic nucleotide-binding domain determines the activation kinetics of SthK. We propose that SthK exists in two forms: trans Pro300 SthK with high ligand binding affinity and fast activation, and cis Pro300 SthK with low affinity and slow activation. Following channel activation, the cis/trans equilibrium, catalyzed by prolyl isomerases, is shifted towards trans, while steady-state channel activity is unaffected. Our results reveal prolyl isomerization as a regulatory mechanism for SthK, and potentially eukaryotic HCN channels. This mechanism could contribute to electrical rhythmicity in cells.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Bacterial Proteins / chemistry
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism
  • Binding Sites
  • Cryoelectron Microscopy
  • Cyclic AMP / metabolism
  • Cyclic Nucleotide-Gated Cation Channels / chemistry*
  • Cyclic Nucleotide-Gated Cation Channels / genetics
  • Cyclic Nucleotide-Gated Cation Channels / metabolism*
  • Cyclosporine / pharmacology
  • Ion Channel Gating / physiology
  • Isomerism
  • Kinetics
  • Models, Molecular
  • Peptidylprolyl Isomerase / metabolism
  • Proline / metabolism
  • Spirochaeta / metabolism*


  • Bacterial Proteins
  • Cyclic Nucleotide-Gated Cation Channels
  • Cyclosporine
  • Proline
  • Cyclic AMP
  • Peptidylprolyl Isomerase

Supplementary concepts

  • Spirochaeta thermophila