A conformational change in the junctional foot protein is involved in the regulation of Ca2+ release from sarcoplasmic reticulum. Studies on polylysine-induced Ca2+ release.
We investigated both conformational changes in the junctional foot protein (JFP) and Ca2+ release from sarcoplasmic reticulum (SR) in parallel after stimulation of triadic vesicles by the JFP-specific ligand, polylysine. To monitor protein conformational change, the JFP was labeled in a site-directed fashion with the fluorescent conformational probe methylcoumarin acetate (MCA) (Kang, J. J., Tarcsafalvi, A., Carlos, A. D., Fujimoto, E., Shahrokh, Z., Thevenin, B. J.-M., Shohet, S. B., and Ikemoto, N. (1992) Biochemistry 31, 3288-3293). The induction of SR Ca2+ release by polylysine produced a rapid increase in the fluorescence intensity of the JFP-bound MCA. The polylysine concentration dependence of the fluorescence change was essentially the same as that of Ca2+ release, suggesting that the two events are tightly coupled. However, the rate constant of MCA fluorescence change was much larger than that of Ca2+ release; i.e. the conformational change preceded Ca2+ release. Prevention of protein conformational change by lysine (0.2 M) inhibited Ca2+ release from SR. Inhibition of Ca2+ release by Mg2+ (5 mM), however, had little effect on the conformational change. These results suggest that binding of polylysine to the JFP produces conformational changes in the protein, which in turn activates the Ca2+ channel, leading to Ca2+ release from the SR.
el-Hayek
R
R
Boston Biomedical Research Institute, Massachusetts 02114, USA.
Yano
M
M
Ikemoto
N
N
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000737:chemistry*; D005453:Fluorescence; D008274:Magnesium / Q000494:pharmacology; D009124:Muscle Proteins / Q000737:chemistry*; D011107:Polylysine / Q000494:pharmacology*; D011487:Protein Conformation; D019837:Ryanodine Receptor Calcium Release Channel; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*
D015220:Calcium Channels; D009124:Muscle Proteins; D019837:Ryanodine Receptor Calcium Release Channel; D011107:Polylysine; D008274:Magnesium; D002118:Calcium