Conformational change of the foot protein of sarcoplasmic reticulum as an initial event of calcium release

Heavy sarcoplasmic reticulum vesicles were labeled with the thiol-reacting fluorescent probe N-(7-dimethylamino-4-methyl-4-coumarinyl)maleimide (DACM), and the DACM-labeled foot protein moiety was purified. The fluorescence intensity of the DACM attached to the foot protein decreased by the addition of low (activating) concentrations of ryanodine, while it increased at higher (inhibitory) concentrations, suggesting that the lower fluorescence represents the active state of the foot protein, while the higher fluorescence, its inactive state. Under conditions that induce Ca²⁺ release from SR (Ca²⁺ jump, addition of Ca²⁺ release inducing reagents such as caffeine and polylysine), the fluorescence intensity of the protein-attached DACM decreased rapidly (e.g. k {reversed tilde equals}7O s^-1 under optimum conditions). The initial rate of Ca²⁺ release from the DACM-labeled SR showed a close correlation with the amplitude of the fluorescence change of the foot protein-attached DACM under variety of conditions; e.g. in the presence of Ca²⁺, polylysine, ATP, and ruthenium red, etc. The fluorescence change of the foot protein was much faster than Ca²⁺ release from SR under a variety of conditions of Ca²⁺ release. We propose that the binding of release triggering reagents to the foot protein induces a rapid conformational change, which in turn regulates Ca²⁺ release.