KINETICS OF RAPID CA2+ RELEASE BY SARCOPLASMIC-RETICULUM - EFFECTS OF CA2+, MG2+, AND ADENINE-NUCLEOTIDES
KINETICS OF RAPID CA2+ RELEASE BY SARCOPLASMIC-RETICULUM - EFFECTS OF CA2+, MG2+, AND ADENINE-NUCLEOTIDES
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DOI：
10.1021/bi00349a033
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发表时间：
1986-01-14
期刊：
影响因子：
2.9
通讯作者：
EVELETH, J
中科院分区：
文献类型：
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作者：
MEISSNER, G;DARLING, E;EVELETH, J
用放射性同位素通量快速猝灭微孔过滤法测定了Ca ~（2+）、腺嘌呤核苷酸和Mg ~（2+）对“重质”肌浆网（SR）囊泡Ca ~（2+）释放行为的影响。从被动加载囊泡的快速45 Ca 2+流出被阻断通过添加Mg 2+和钌红。在pH 7和10-9 M Ca ~（2+）条件下，囊泡释放~（45）C ~（2+）的速率较低（k = 0.1 s ~（-1））。将外部Ca 2+浓度增加至4 μ M或加入5 mM ATP或ATP类似物腺苷5“-（β，γ- AMP-PCP的45 Ca ~（2+）释放速率居中。在含有4 μ M Ca 2+和5 mM AMP-PCP的培养基中观察到最大释放速率，并且具有30-100 s-1的一级速率常数。Mg ~（2+）部分抑制Ca ~（2+）和核苷酸诱导的~（45）Ca ~（2+）外排。在不存在AMP-PCP的情况下，在5 mM Mg 2+或5 mM Ca 2+下，45 Ca 2+释放被完全抑制。系统地改变释放介质的组成，并且以Hill方程的形式表示通量数据。ATP和AMP-PCP激活Ca ~（2+）释放的表观n值为1.6 ~ 1.9。Ca ~（2+）激活的Hill系数（n = 0.8-2.1）与核苷酸和Mg ~（2+）浓度有关，而Mg ~（2+）抑制的Hill系数（n = 1.1-1.6）则与外源Ca ~（2+）浓度有关。这些结果表明，重SR囊泡含有一个“钙释放通道”，这是能够进行钙离子的速率与那些发现在完整的肌肉。Ca ~（2+）、AMP-PCP（ATP）和Mg ~（2+）似乎作用于通道的非相互作用或相互作用位点。
A radioisotope flux-rapid-quench-Millipore filtration method is described for determining the effects of Ca2+, adenine nucleotides, and Mg2+ on the Ca2+ release behavior of "heavy" sarcoplasmic reticulum (SR) vesicles. Rapid 45Ca2+ efflux from passively loaded vesicles was blocked by the addition of Mg2+ and ruthenium red. At pH 7 and 10-9 M Ca2+, vesicles released 45C2+ with a low rate (k = 0.1 s-1). An increase in external Ca2+ concentration to 4 .mu.M or the addition of 5 mM ATP or the ATP analogue adenosine 5''-(.beta., .gamma.-methylenetriphosphate) (AMP-PCP) resulted in intermediate 45Ca2+ release rates. The maximal release rate was observed in media containing 4 .mu.M Ca2+ and 5 mM AMP-PCP and had a first-order rate constant of 30-100 s-1. Mg2+ partially inhibited Ca2+- and nucleotide-induced 45Ca2+ efflux. In the absence of AMP-PCP, 45Ca2+ release was fully inhibited at 5 mM Mg2+ or 5 mM Ca2+. The composition of the release media was systematically varied, and the flux data were expressed in the form of Hill equations. The apparent n values of activation of Ca2+ release by ATP and AMP-PCP were 1.6-1.9. The Hill coefficient of Ca2+ activation (n = 0.8-2.1) was dependent on nucleotide and Mg2+ concentrations, whereas the one of Mg2+ inhibition (n = 1.1-1.6) varied with external Ca2+ concentration. These results suggest that heavy SR vesicles contain a "Ca2+ release channel" which is capable of conducting Ca2+ at rates comparable with those found in intact muscle. Ca2+, AMP-PCP (ATP), and Mg2+ appear to act at noninteracting or interacting sites of the channel.