Selective modification of functionally distinct sulfhydryl groups of sarcoplasmic reticulum Ca2+,Mg2+-adenosine triphosphatase with N-ethylmaleimide.
Selective modification of functionally distinct sulfhydryl groups of sarcoplasmic reticulum Ca2+,Mg2+-adenosine triphosphatase with N-ethylmaleimide.
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用 N-乙基马来酰亚胺选择性修饰肌浆网 Ca2、Mg2-腺苷三磷酸酶的功能不同的巯基。
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发表时间：
1980
期刊：
影响因子：
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通讯作者：
Y. Kaziro
中科院分区：
文献类型：
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作者：
M. Kawakita;K. Yasuoka;Y. Kaziro
用N-乙基马来酰亚胺（NEM）在pH 7.0下处理兔肌浆网囊泡。在1.5mM NEM下，在30 ℃下25分钟内每摩尔ATP酶肽仅修饰4个SH基团。其中两个是Ca ~（2+）转运所必需的，一个参与E-P形成（SHF），另一个参与其分解（SHD），而另外两个显然是非必需的（SHN和SHN '）。首先修改SHN，然后按顺序修改SHD、SHN '和SHF。SHD的修饰伴随着Ca ~（2+）转运活性的丧失，而E-P形成活性存活，直到最小反应性的（SHF）被修饰。在较低的NEM浓度（4 × 10（-5）M）下，SHN可以被选择性地修饰而不损失酶活性。在Ca 2+离子存在下，SHF可以被腺苷-5 '-基-亚氨二磷酸（AMP-P（NH）P）保护，而SHD则不能。SHD在无Ca 2+（小于10（-7）M）时的反应性明显低于有Ca 2+时。这些SH基团的反应性的变化可能与ATP酶分子的构象变化引起的Ca ~（2+）和ATP的结合。
Rabbit sarcoplasmic reticulum vesicles were treated with N-ethylmaleimide (NEM) at pH 7.0. At 1.5 mM NEM, only 4 SH groups per mol of ATPase peptide were modified in 25 min at 30 degrees C. Two of these are essential for Ca2+ transport, one being involved in E-P formation (SHF), and the other in its decomposition (SHD), whereas the other two are apparently non-essential (SHN and SHN'). SHN was modified first, followed by SHD, SHN', and SHF, in this order. Modification of SHD was accompanied by the loss of Ca2+-transport activity, while E-P forming activity survived until the least reactive one (SHF) was modified. At a lower NEM concentration (4 x 10(-5) M) SHN could be selectively modified without loss of the enzyme activity. SHF could be protected by adenyl-5'-yl-imidodiphosphate (AMP-P(NH)P) in the presence of Ca2+ ions, whereas SHD was not. SHD was distinctly less reactive in the absence of Ca2+ (less than 10(-7) M) than in its presence. Changes in the reactivity of these SH groups may be related to conformational changes of the ATPase molecule induced by the binding of Ca2+ and ATP.