Ca2+-myristoyl switch and membrane binding of chemically acylated neurocalcins
Ca2+-myristoyl switch and membrane binding of chemically acylated neurocalcins
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DOI：
10.1021/bi010188e
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发表时间：
2001-07-10
期刊：
影响因子：
2.9
通讯作者：
Chopineau, J
中科院分区：
文献类型：
--
作者：
Béven, L;Adenier, H;Chopineau, J
Neurocalin是一个新的神经细胞钙感受器家族的成员，属于EF-Hand钙结合蛋白超家族。神经降钙素在体内的N-末端发生肉豆蔻酰化，并能以钙和肉豆蔻基依赖的方式与生物膜结合。这一被称为“钙-肉豆蔻基开关”的过程对光感受器特定蛋白Recovery in以及其他几种神经元钙传感器有最好的描述。在这里，我们使用反胶束在其N-端用不同长度的脂肪酸(从C12到C16)化学酰化非肉豆蔻酰化的神经钙蛋白。这种方法使我们能够制备神经钙素衍生物，其中单一脂肪酸通过酰胺键选择性地连接到多肽链的N-末端甘氨酸。然后通过与磷脂小泡共沉积和表面等离子体共振光谱(Biacore)直接与脂单分子层结合来检测单酰化神经钙蛋白的膜结合特性。我们的结果表明，用月桂酸、肉豆蔻酸或棕榈酸单酰化的神经钙素能够以钙依赖的方式与膜结合。这表明，钙-肉豆蔻酰基开关可以与不同的脂质部分一起发挥作用，而不是严格地限制在肉豆蔻酸中。随意修饰连接到N-末端甘氨酸的脂肪酸的能力对于分析脂肪酸部分对这类神经元钙传感器的生物功能的贡献应该是有用的，
Neurocalcin is a member of a novel family of neuronal calcium sensors that belongs to the superfamily of EF-hand Ca2+-binding proteins. Neurocalcin is myristoylated on its N-terminus in vivo and can associate with biological membranes in a calcium and myristoyl-dependent manner. This process known as "Ca2+-myristoyl switch" has been best described for the photoreceptor specific protein, recoverin, as well as for several other neuronal calcium sensors. Here, we used reversed micelles to chemically acylate nonmyristoylated neurocalcin at its N-terminus with fatty acids of different lengths (from C12 to C16). This approach allowed us to prepare neurocalcin derivatives in which a single fatty acid is selectively linked to the N-terminal glycine of the polypeptide chain through an amide bond. The membrane binding properties of the monoacylated neurocalcins were then examined by cosedimentation with phospholipid vesicles and direct binding to lipid monolayers by surface plasmon resonance spectroscopy (Biacore). Our results show that neurocalcins monoacylated with lauric, myristic, or palmitic acid were able to associate with membrane in a calcium-dependent manner. This indicates that the Ca2+-myristoyl switch can function with different lipid moieties and is not strictly restricted to myristate. The ability to modify at will the fatty acid linked to the N-terminal glycine should be useful to analyze the contribution of the fatty acid moiety to the biological function of this family of neuronal calcium sensors,