Biochemical characterization of copine:: A ubiquitous Ca2+-dependent, phospholipid-binding protein
Biochemical characterization of copine:: A ubiquitous Ca2+-dependent, phospholipid-binding protein
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DOI：
10.1021/bi0019949
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发表时间：
2000-12-26
期刊：
影响因子：
2.9
通讯作者：
Creutz, CE
中科院分区：
文献类型：
--
作者：
Tomsig, JL;Creutz, CE
copines是一组新的Ca 2+依赖性磷脂结合蛋白，首先从四脲草履虫（Paramecium tetraurelia）[Creutz，C. E、等人（1998）J.Biol.Chem.273，1393-1402]，并且在从植物到人类的广泛生物体中发现。它们具有由两个C2结构域组成的Ca 2+和磷脂结合结构域和在C-末端部分的核心结构域，其与在某些整联蛋白中发现的A结构域同源。我们在这里提供的第一个描述的性质和分布的天然哺乳动物copine，copine I。这种蛋白质在所有主要的成年大鼠器官中表达，如通过用anticopine抗体探测大鼠器官匀浆的Western印迹所证明的。在脾脏中发现最高水平的Copine。本文报道了从牛脾中纯化Copine的方法。纯化的天然Copine是一种58 kDa的单体，其表现出Ca 2+自缔合以形成高阶多聚体，以及Ca 2+依赖性磷脂结合活性，相对于中性磷脂，对带负电荷的磷脂具有优先性，并且相对于Mg 2+，对Ca 2+具有选择性。与富含磷脂酰丝氨酸的囊泡的半最大结合发生在Ca 2+浓度为1 - 10 μ M的Copine I，其表现出Mn 2+结合活性，该活性被Mg 2+强烈竞争并被Ca 2+部分竞争，这表明Copine IA结构域可能是与整联蛋白中发现的相似的功能性MIDAS金属结合位点[Lee，J.O.，等人（1995）Cell 80，631-638]。Copine在结合膜和靶蛋白或小分子中的作用进行了讨论。
The copines are a novel group of Ca2+-dependent, phospholipid-binding proteins first isolated from Paramecium tetraurelia [Creutz, C. E., et al. (1998) J. Biol. Chem. 273, 1393-1402] and found in a wide range of organisms, from plants to humans. They have a Ca2+ and phospholipid-binding domain consisting of two C2 domains and a core domain in the C-terminal portion that is homologous to the A domain found in certain integrins. We provide here the first description of the properties and distribution of a native mammalian copine, copine I. This protein is expressed in all major adult rat organs as demonstrated by probing Western blots of rat organ homogenates with anticopine antibodies. The highest levels of copine are found in the spleen. A protocol for purifying copine to homogeneity from bovine spleen is described. Purified native copine is a 58 kDa monomer that exhibits Ca2+ self-association to form higher-order multimers, and Ca2+-dependent, phospholipid binding activity with preference for negatively charged phospholipids over neutral phospholipids and selectivity for Ca2+ over Mg2+. Half-maximal association with vesicles enriched in phosphatidylserine occurs at Ca2+ concentrations between 1 and 10 muM Copine I exhibits Mn2+ binding activity that is strongly competed by Mg2+ and partially competed by Ca2+, suggesting that the copine I A domain may be a functional MIDAS metal binding site similar to that found in integrins [Lee, J. O., et al. (1995) Cell 80, 631-638]. Roles for copine in binding membranes and target proteins or small molecules are discussed.