Shoshan V, MacLennan D H, Wood D S
Proc Natl Acad Sci U S A. 1981 Aug;78(8):4828-32. doi: 10.1073/pnas.78.8.4828.
Sarcoplasmic reticulum vesicles from mammalian skeletal muscle have previously been shown to develop a proton gradient (alkaline inside) of 0.15-0.5 pH units during active Ca2+ uptake. We found that dissipation of this gradient by the proton ionophores gramicidin, nigericin, and carbonyl cyanide p-trichloromethoxyphenylhydrazone caused a rapid transient tension in skinned rabbit psoas muscle fibers. Increases, but not decreases, in medium pH of approximately 0.2 units over the range from pH 6.5 to pH 7.5 also elicited transient tensions. In isolated vesicles, physiological levels of Ca2+ (3.3 microM), inhibited pH-induced Ca2+ release. Dicyclohexylcarbodiimide blocked pH- and ionophore-induced Ca2+ release under conditions in which it could bind to sarcoplasmic reticulum proteins but did not inhibit Ca2+ uptake. We propose that a proton gradient generated across sarcoplasmic reticulum membranes during Ca2+ uptake maintains a Ca2+ release channel in a closed conformation and that dissipation of this gradient permits the Ca2+ release channel to open. We further propose that elevated myoplasmic Ca2+ also causes the Ca2+ channel to close, permitting Ca2+ uptake through Ca2+/Mg2+-ATPase to function effectively. As the proteolipids of sarcoplasmic reticulum bind dicyclohexylcarbodiimide under conditions in which Ca2+ release is blocked and as they have previously been shown to have Ca2+ ionophoric activity, we propose that the Ca2+-release channel either resides in the proteolipids or is controlled by H+ fluxes through the proteolipids.
先前已表明，来自哺乳动物骨骼肌的肌浆网囊泡在主动摄取Ca2+过程中会形成0.15 - 0.5个pH单位的质子梯度（内部呈碱性）。我们发现，质子离子载体短杆菌肽、尼日利亚菌素和羰基氰对氯苯腙消除这种梯度会在去表皮的兔腰大肌纤维中引起快速短暂的张力。在pH 6.5至pH 7.5范围内，培养基pH值升高约0.2个单位（而非降低）也会引发短暂张力。在分离的囊泡中，生理水平的Ca2+（3.3 microM）会抑制pH诱导的Ca2+释放。二环己基碳二亚胺在能够与肌浆网蛋白结合的条件下，阻断了pH和离子载体诱导的Ca2+释放，但不抑制Ca2+摄取。我们提出，在Ca2+摄取过程中跨肌浆网膜产生的质子梯度使Ca2+释放通道保持关闭构象，而这种梯度的消除会使Ca2+释放通道打开。我们还提出，肌浆内Ca2+升高也会导致Ca2+通道关闭，从而使通过Ca2+/Mg2+-ATP酶的Ca2+摄取能够有效发挥作用。由于肌浆网的蛋白脂质在Ca2+释放被阻断的条件下会结合二环己基碳二亚胺，且先前已表明它们具有Ca2+离子载体活性，我们提出Ca2+释放通道要么存在于蛋白脂质中，要么受通过蛋白脂质的H+通量控制。
