Functional and Structural Interactions between Ca2+, ATP and Caffeine Binding Sites of Skeletal Muscle Ryanodine Receptor (RyR1)

Type 1 ryanodine receptor (RyR1) releases Ca2+ ions from the sarcoplasmic reticulum of skeletal muscle to initiate contraction. Multiple endogenous and exogenous effectors activate RyR1. Cryo-electron microscopy has identified binding sites three co-activators Ca2+, ATP caffeine, however, mechanism co-regulation synergy between these activators remains be determined. Here, we tested hypothesis that both caffeine interact with site RyR1 thereby sensitize channel activation by Ca2+. A [3H]ryanodine ligand assay computational methods using cryo-EM micrograph densities determined regulation corresponding structural changes in site. We report AMPPCP absence presence a further increase occurs caffeine. Computational analysis suggested modulate conformation through polar interactions central terminal domain. In Ca2+-only bound RyR1, inter-domain crucial for Ca2+-ATP-caffeine communication are predicted. Caffeine variants RyR1-W4716R -I4996A RyR1-M4954A K4211M/R4215F provide additional evidence interaction regulate conclude ATP, network allosteric interactions. Supported NIH grants AR18687 HL147054.