Ca²⁺ inactivation of the mammalian ryanodine receptor type 1 in a lipidic environment revealed by cryo-EM.

Abstract

Activation of the intracellular Ca²⁺ channel ryanodine receptor (RyR) triggers a cytosolic Ca²⁺ surge, while elevated cytosolic Ca²⁺ inhibits the channel in a negative feedback mechanism. Cryogenic electron microscopy of rabbit RyR1 embedded in nanodiscs under partially inactivating Ca²⁺ conditions revealed an open and a closed-inactivated conformation. Ca²⁺ binding to the high-affinity site engages the central and C-terminal domains into a block, which pries the S6 four-helix bundle open. Further rotation of this block pushes S6 toward the central axis, closing (inactivating) the channel. Main characteristics of the Ca²⁺-inactivated conformation are downward conformation of the cytoplasmic assembly and tightly knit subunit interface contributed by a fully occupied Ca²⁺ activation site, two inter-subunit resolved lipids, and two salt bridges between the EF hand domain and the S2-S3 loop validated by disease-causing mutations. The structural insight illustrates the prior Ca²⁺ activation prerequisite for Ca²⁺ inactivation and provides for a seamless transition from inactivated to closed conformations.