DETECTION OF CALCIUM-BINDING PROTEINS BY CA-45 AUTORADIOGRAPHY ON NITROCELLULOSE MEMBRANE AFTER SODIUM DODECYL-SULFATE GEL-ELECTROPHORESIS
DETECTION OF CALCIUM-BINDING PROTEINS BY CA-45 AUTORADIOGRAPHY ON NITROCELLULOSE MEMBRANE AFTER SODIUM DODECYL-SULFATE GEL-ELECTROPHORESIS
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DOI：
10.1093/oxfordjournals.jbchem.a134633
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发表时间：
1984-01-01
影响因子：
2.7
通讯作者：
EBASHI, S
中科院分区：
文献类型：
--
作者：
MARUYAMA, K;MIKAWA, T;EBASHI, S
本文介绍了一种检测混合蛋白中钙结合蛋白的新方法。将可能包含Ca结合蛋白的样品进行SDS[十二烷基硫酸钠]-聚丙烯酰胺凝胶电泳，然后电泳转移到硝酸纤维素膜上。然后将膜与45 Ca孵育，以通过放射自显影法检测作为放射性条带的Ca结合蛋白。纯化的肌钙蛋白-C、钙调蛋白、肌球蛋白DTNB [5，5“-二硫代双（2-硝基苯甲酸）] L链和小清蛋白通过该方法被清楚地鉴定。在鸡骨骼肌全匀浆中，肌球蛋白DTNB轻链、肌钙蛋白C和55 K [千] Ca结合蛋白具有放射性。在蛙骨骼肌中，约13- 15 K和70 K的小分子量蛋白质似乎是钙结合蛋白。在鲤鱼骨骼肌中，小分子量蛋白包括小清蛋白和2个约80 K的蛋白质似乎与Ca ~（2+）结合。扇贝横纹肌中存在两种高分子量钙结合蛋白。该过程可以在24小时内完成，并且可以在起始样品中检测到少至2 μ g的Ca结合蛋白。在适当的条件下，有可能只检测高亲和力的钙结合蛋白。
A new simple method of detecting Ca binding proteins in a protein mixture is described. A sample which might include Ca binding proteins was subjected to SDS[sodium dodecylsulfate]-polyacrylamide gel electrophoresis and then electrophoretically transferred to a nitrocellulose membrane. The membrane was then incubated with 45Ca to detect Ca-binding proteins as radioactive bands by autoradiography. Purified troponin-C, calmodulin, myosin DTNB [5,5''-dithiobis(2-nitrobenzoic acid)] L chain and parvalbumin were clearly identified by this method. In the whole homogenate of chicken skeletal muscle, myosin DTNB light chain, troponin-C and 55K [thousand] Ca binding protein were found to be radioactive. In frog skeletal muscle, small MW proteins of approximately 13-15K and 70K protein appeared to be the Ca binding proteins. In carp skeletal muscle, small MW proteins including parvalbumin and 2 proteins of about 80K seemed to bind Ca2+. Two high MW Ca binding proteins were present in scallop striated muscle. The procedure can be completed within 24 h and can detect as little as 2 .mu.g of Ca binding protein in the starting sample. Under appropriate conditions, it was possible to detect only high affinity Ca binding proteins.