Luminal pH regulated calcium release kinetics in sarcoplasmic reticulum vesicles.
Calcium binding to triads isolated from rabbit skeletal muscle followed a single hyperbolic function in the pH range 5.5-8.0. Maximal binding was obtained at pH 8.0; decreasing the pH decreased the binding capacity and, at pH < or = 6.0, increased Kd 2-fold. These results indicate that lowering the pH diminished calcium binding to calsequestrin, since this protein is the primary source of calcium binding sites in triads. Luminal pH had a marked effect on calcium release induced by 2 mM ATP, at pCa 5.0, pH 6.8. At a constant luminal [Ca2+] of 0.1 mM, release rate constants (k) and initial rates of release increased steadily as a function of decreasing luminal pH; at luminal pH 7.5, values of k < 0.4 s-1 were found, whereas at pH 5.5 values of k approximately 10 S-1 were obtained. Increasing luminal [Ca2+] from 0.05 mM to 0.7 mM had no effect on the k values measured at luminal pH 5.5. In contrast, at pH 6.8, increasing luminal [Ca2+] produced a marked increase in k values, that reached maximal values of k approximately 10 S-1 at 0.7 mM luminal [Ca2+]. Control experiments using fluorescent pH indicators showed that luminal pH did not change significantly during calcium release. It is proposed that luminal protons or calcium induces conformational changes in calsequestrin that in turn promote activation of the calcium release channels.
Donoso
P
P
Departamento de Fisiología y Biofísica, Facultad de Medicina, Universidad de Chile, Santiago, Chile. pdonoso@machi.med.ucbile.cl
Beltrán
M
M
Hidalgo
C
C
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000378:metabolism; D002155:Calsequestrin / Q000737:chemistry / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D007700:Kinetics; D009124:Muscle Proteins / Q000378:metabolism; D018482:Muscle, Skeletal / Q000378:metabolism; D011487:Protein Conformation; D011522:Protons; D011817:Rabbits; D019837:Ryanodine Receptor Calcium Release Channel; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*
D015220:Calcium Channels; D002155:Calsequestrin; D009124:Muscle Proteins; D011522:Protons; D019837:Ryanodine Receptor Calcium Release Channel; D002118:Calcium