ISOLATION AND CHARACTERIZATION OF MUSCLE MEMBRANES USING SURFACE-SPECIFIC LABELS
ISOLATION AND CHARACTERIZATION OF MUSCLE MEMBRANES USING SURFACE-SPECIFIC LABELS
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DOI：
10.1073/pnas.74.1.34
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发表时间：
1977-01-01
影响因子：
11.1
通讯作者：
WONG, M
中科院分区：
文献类型：
--
作者：
BARCHI, RL;BONILLA, E;WONG, M
通过差速离心法从大鼠肌肉中纯化膜，该方法避免使用盐提取或在升高的温度下孵育。在连续蔗糖梯度中，膜限制性囊泡的三个群体被定义为具有1.112（级分I）、1.141（级分II）和1.158（级分III）g/ml的平均密度。乳过氧化物酶催化的碘化的完整的肌肉分离膜之前，导致在馏分I中的最高比活性，虽然所有馏分分离后同样标记。小麦胚芽凝集素（125 I标记）在低浓度下与完整肌肉优先标记的组分I孵育。对先前分离的组分的平行研究表明，组分I也含有最高浓度的麦胚凝集素的潜在受体。在整个肌肉的实验中，伴刀豆球蛋白A结合主要是肌膜与轻微的变量结合T-管和核膜，但没有与分离的膜片段的结合研究表明重结合伴刀豆球蛋白A的膜在馏分I与分散的结合在馏分II和III。Na+K+ Mg 2 + ATP酶在组分I中特异性富集，但也以类似于125 I标记的比例存在于组分II中。Ca ~（2+）ATP酶在组分II中最活跃，但在组分I中存在显著水平。组分I主要含有肌膜，而T-管膜代表组分II的重要组成部分。组分I的Ca ~（2+）ATP酶活性是肌膜固有的。
Membranes were purified from rat muscle by a differential centrifugation method that avoids the use of salt extraction or incubations at elevated temperature. Three populations of membrane-limited vesicles were defined having average densities of 1.112 (fraction I), 1.141 (fraction II) and 1.158 (fraction III) g/ml in a continuous sucrose gradient. Lactoperoxidase-catalyzed iodination of intact muscle prior to isolation of membranes resulted in highest specific activity in fraction I, although all fractions were equally labeled after isolation. Wheat germ agglutinin (125I-labeled) incubated at low concentration with intact muscle preferentially labeled fraction I. Parallel studies on previously isolated fractions indicated that fraction I also contained the highest concentration of potential receptors for wheat germ agglutinin. In experiments on whole muscle, concanavalin A bound predominantly to sarcolemma with slight variable binding to T-tubular and nuclear membrane but no binding studies with isolated membrane fragments indicated heavy binding of concanavalin A by membranes in fraction I with scattered binding in fractions II and III. Na+K+Mg2+ ATPase was specifically enriched in fraction I but was also present in fraction II in a proportion similar to 125I labeling. Ca2+ ATPase was most active in fraction II but present in significant levels in fraction I. Fraction I contained predominantly sarcolemma membrane, while T-tubular membrane represented a significant component of fraction II. Ca2+ ATPase activity in fraction I was intrinsic to the sarcolemma.