Abstract

Ryanodine receptors (RyRs) form calcium release channels located in the membranes of the sarcoplasmic and endoplasmic reticulum. RyRs play a major role in excitation-contraction coupling and other Ca2+-dependent signalling events, and consist of several globular domains that together form a large assembly. Here we describe the crystal structures of the SPRY1 and tandem-repeat domains at 1.2–1.5 Å resolution, which reveal several structural elements not detected in recent cryo-EM reconstructions of RyRs. The cryo-EM studies disagree on the position of SPRY domains, which had been proposed based on homology modelling. Computational docking of the crystal structures, combined with FRET studies, show that the SPRY1 domain is located next to FK506-binding protein (FKBP). Molecular dynamics flexible fitting and mutagenesis experiments suggest a hydrophobic cluster within SPRY1 that is crucial for FKBP binding. A RyR1 disease mutation, N760D, appears to directly impact FKBP binding through interfering with SPRY1 folding.

Highlights

  • Ryanodine receptors (RyRs) form calcium release channels located in the membranes of the sarcoplasmic and endoplasmic reticulum

  • Several studies have implied that unbinding of FKBP12 or FKBP12.6 may contribute to diseases associated with RyR Ca2 þ leak like muscular dystrophy, sarcopenia, heart failure, diabetes, or Alzheimer’s21–24

  • These discrepancies cannot be ascribed to different isoforms, because the SPRY1 sequence is highly conserved among all three RyR isoforms (Fig. 2d)

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Summary

Introduction

Ryanodine receptors (RyRs) form calcium release channels located in the membranes of the sarcoplasmic and endoplasmic reticulum. Computational docking of the crystal structures, combined with FRET studies, show that the SPRY1 domain is located next to FK506-binding protein (FKBP). Lowresolution cryo-EM studies[27,28,29], as well as fluorescence resonance energy transfer (FRET) measurements[30] show that FKBP12 and FKBP12.6 bind in the same position and orientation to the periphery of the cytosolic cap, at the junction between the ‘clamp’ and ‘handle’ domains. This binding mode has been confirmed with the higher resolution cryo-EM studies, which showed either FKBP12 The exact binding determinants for FKBPs within the RyR protein sequence have remained ambiguous, as numerous sites have been identified throughout the channel[31,32,33,34]

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