In silico assessment of the conduction mechanism of the Ryanodine Receptor 1 reveals previously unknown exit pathways

Abstract

The ryanodine receptor 1 is a large calcium ion channel found in mammalian skeletal muscle. The ion channel gained a lot of attention recently, after multiple independent authors published near-atomic cryo electron microscopy data. Taking advantage of the unprecedented quality of structural data, we performed molecular dynamics simulations on the entire ion channel as well as on a reduced model. We calculated potentials of mean force for Ba2+, Ca2+, Mg2+, K+, Na+ and Cl− ions using umbrella sampling to identify the key residues involved in ion permeation. We found two main binding sites for the cations, whereas the channel is strongly repulsive for chloride ions. Furthermore, the data is consistent with the model that the receptor achieves its ion selectivity by over-affinity for divalent cations in a calcium-block-like fashion. We reproduced the experimental conductance for potassium ions in permeation simulations with applied voltage. The analysis of the permeation paths shows that ions exit the pore via multiple pathways, which we suggest to be related to the experimental observation of different subconducting states.

The Ca2+ Permeation Mechanism of the Ryanodine Receptor Revealed by a Multi-Site Ion Model

“…By calculating the residence time of carboxylate oxygen of negatively charged residues within a sphere with a radius of 5 Å around the binding site L, we identified that the binding site L is formed by the interaction of Ca 2+ ions with D4899, E4900, and D4903 residues ( Fig. 3b), which agrees well with previous experimental and computational studies 28,30 . The large probability of finding Ca 2+ in the selectivity filter also indicates that the Ca 2+ can easily accumulate around the luminal vestibule and move into the upper filter, and the rate-determining step of permeation is the process of passing through the lower selectivity filter or gate constrictions.…”

“…2c). Since the channel is in its open state, this indicated that the default Ca 2+ model gave us qualitatively wrong simulation results here, as also observed by another recent study 28 . A close inspection of the MD trajectories showed that the Ca 2+ ions were tightly bound to the protein, again confirming that the binding affinity between Ca 2+ and protein is too strong with the conventional Ca 2+ model.…”

“…The interaction between Ca 2+ and protein is of great importance in studying Ca 2+ The ion binding and permeation mechanisms have been widely studied for K + and Na + channels with molecular dynamics simulations 10-12,14-17 , but only rarely studied for Ca 2+ channels with conventional ion models 28 . From our MD simulations with the new multi-site Ca 2+ model, the major Ca 2+ binding sites in the open-state RyR1 were determined to be near the D4900 and G4894 residues on the luminal side, and the rate-determining step of permeation is found to be the step of passing through the selectivity filter constriction, where Ca 2+ dwell for a relatively long time before passing through ( Fig.…”

“…The conventional ion models generally work well in studying ion-protein interactions for K + or Na + , but fail consistently for Ca 2+ ions 9 , and therefore no calcium permeation was observed in previous MD studies of Ca 2+ channels 28 . We performed MD simulations on the type-1 ryanodine receptor (RyR1), an intracellular calcium release channel required for skeletal muscle contraction, with our Ca 2+ model.…”

Correlation of phenotype with genotype and protein structure in RYR1-related disorders

“…Many variants within the CS plane of interest occur at positions that are exposed to solution, often in open cavities, suggesting these cavities have important roles in channel function. Indeed, recent biophysical simulation studies have revealed additional ion conduction pathways that permeate the lateral aspect of the RyR1 CS and may thereby enable lateral Ca 2+ efflux into the cytoplasm [22]. …”