Cardiac ryanodine receptor: Selectivity for alkaline earth metal cations points to the EF-hand nature of luminal binding sites

Bioelectrochemistry. 2016 Jun:109:49-56. doi: 10.1016/j.bioelechem.2016.01.002. Epub 2016 Jan 28.


A growing body of evidence suggests that the regulation of cardiac ryanodine receptor (RYR2) by luminal Ca(2+) is mediated by luminal binding sites located on the RYR2 channel itself and/or its auxiliary protein, calsequestrin. The localization and structure of RYR2-resident binding sites are not known because of the lack of a high-resolution structure of RYR2 luminal regions. To obtain the first structural insight, we probed the RYR2 luminal face stripped of calsequestrin by alkaline earth metal divalents (M(2+): Mg(2+), Ca(2+), Sr(2+) or Ba(2+)). We show that the RYR2 response to caffeine at the single-channel level is significantly modified by the nature of luminal M(2+). Moreover, we performed competition experiments by varying the concentration of luminal M(2+) (Mg(2+), Sr(2+) or Ba(2+)) from 8 mM to 53 mM and investigated its ability to compete with 1mM luminal Ca(2+). We demonstrate that all tested M(2+) bind to exactly the same RYR2 luminal binding sites. Their affinities decrease in the order: Ca(2+)>Sr(2+)>Mg(2+)~Ba(2+), showing a strong correlation with the M(2+) affinity of the EF-hand motif. This indicates that the RYR2 luminal binding regions and the EF-hand motif likely share some structural similarities because the structure ties directly to the function.

Keywords: Alkaline earth metal divalent; Cardiac muscle; Luminal regulation; Planar lipid bilayer; Ryanodine receptor.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Anticonvulsants / metabolism
  • Anticonvulsants / pharmacology*
  • Barium / metabolism
  • Binding Sites
  • Caffeine / pharmacology
  • Calcium / metabolism*
  • Cations, Divalent / metabolism
  • Central Nervous System Stimulants / pharmacology
  • EF Hand Motifs
  • Humans
  • Magnesium / metabolism
  • Metals, Alkaline Earth / metabolism*
  • Molecular Sequence Data
  • Myocardium / chemistry
  • Myocardium / metabolism*
  • Phenobarbital / metabolism
  • Phenobarbital / pharmacology*
  • Ryanodine Receptor Calcium Release Channel / chemistry
  • Ryanodine Receptor Calcium Release Channel / metabolism*
  • Sequence Alignment
  • Strontium / metabolism


  • Anticonvulsants
  • Cations, Divalent
  • Central Nervous System Stimulants
  • Metals, Alkaline Earth
  • Ryanodine Receptor Calcium Release Channel
  • Barium
  • Caffeine
  • Magnesium
  • Calcium
  • Phenobarbital
  • Strontium