Determination of the phosphorylation sites of smooth muscle caldesmon by protein kinase C.
Determination of the phosphorylation sites of smooth muscle caldesmon by protein kinase C.
复制标题
蛋白激酶 C 测定平滑肌钙结合蛋白的磷酸化位点。
DOI：
10.1016/0003-9861(91)90232-8
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发表时间：
1991
影响因子：
3.9
通讯作者：
Hornick,T
中科院分区：
文献类型：
--
作者：
Ikebe,M;Hornick,T
平滑肌钙调蛋白被蛋白激酶C磷酸化至1.90 mol P/mol钙调蛋白。磷酸化钙调蛋白被胰蛋白酶完全消化，产生的磷酸肽通过C-8和C-18反相色谱纯化。分离得到四个磷酸肽。这些肽的氨基酸序列进行了测定，并确定了两个磷酸丝氨酸。两者均位于C-末端结构域的丝氨酸-587和丝氨酸-726处。根据磷酸化的时间进程，丝氨酸-587被认为是首选的网站。还检测了蛋白C对钙调蛋白的磷酸化对acto-H-裂肌球蛋白ATP酶活性抑制的影响。未磷酸化的钙调素抑制ATP酶活性达60%，而磷酸化的钙调素几乎不抑制ATP酶活性。因此，可以得出结论，丝氨酸-726和丝氨酸-587处的磷酸化逆转了钙调蛋白的抑制活性。
Smooth muscle caldesmon was phosphorylated by protein kinase C up to 1.90 mol P/mol caldesmon. Phosphorylated caldesmon was completely digested by trypsin and the produced phosphopeptides were purified by C-8 and C-18 reverse phase chromatography. Four phosphopeptides were isolated. The amino acid sequences of these peptides were determined and two phosphoserines were identified. Both were localized in the C-terminal domain at serine-587 and serine-726. By following the time course of phosphorylation, serine-587 was found to be the preferred site. Effects of the phosphorylation of caldesmon by protein C on the inhibition of acto-H-meromyosin ATPase activity was also examined. While unphosphorylated caldesmon inhibited the ATPase activity by 60%, phosphorylated caldesmon hardly inhibited the ATPase activity. Therefore, it was concluded that the phosphorylation at serine-726 and serine-587 reverses the inhibitory activity of caldesmon.