Calcium Gating by the DHPR-RyR1 Pair

Abstract

In skeletal muscle the L-type voltage-gated calcium channel (DHPR) in the t-tubule coexists in a tight functional interaction with the sarcoplasmic reticulum calcium release channel (RyR1). By means of this interaction a depolarization-induced conformational change in the DHPR is translated into RyR1's opening, and a massive calcium release from intracellular stores. We are performing structural studies on purified DHPRs and RyRs to understand structural details involved in the gating mechanism.The DHPR is a heteropentamer with total molecular weight of ∼450 kDa. Up to now he best structural knowledge has been gained by electron microscopy, although its relative small size has limited the resolution obtained to date. Our new 25A resolution 3D reconstruction shows two distinct parts: a main body shaped like an irregular pentagon with distinct corners, and a hook-shaped feature. Consistent with the considerable conservation of membrane topology among voltage-gated channels, a good part of the main body can be closely fitted with an atomic structure of a full-length potassium channel, and this in turn is helping to locate the RyR1-interacting domains identified using biochemistry and molecular biology techniques.The RyR is a large homotetramer of 2.2 MDa, which has facilitated its structural study by 3D cryo-electron microscopy. Its reproducible 3D structure consists of a large cytoplasmic domain and a smaller transmembrane domain. Our 3D reconstructions of RyR1 in the open and closed states at 10 A resolution show that the ion pathway consists of two right-handed bundles converging into a constriction (putative ion gate) that changes its diameter by ∼4A upon gating. Although the molecular distance between the putative ion gate and the closest site of proximity to the DHPR is very large (>130 A), the conformational changes associated with gating are generalized, suggesting long-range allosteric pathways connecting these distant domains.