Ikebe M, Hornick T
Department of Physiology and Biophysics, Case Western Reserve University, Cleveland, Ohio 44106.
Arch Biochem Biophys. 1991 Aug 1;288(2):538-42. doi: 10.1016/0003-9861(91)90232-8.
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 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.
平滑肌钙调蛋白被蛋白激酶C磷酸化，磷酸化程度高达1.90摩尔磷/摩尔钙调蛋白。磷酸化的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反相色谱法纯化。确定了四个磷酸肽，并鉴定出两个磷酸丝氨酸。两者都位于C末端结构域的丝氨酸-587和丝氨酸-726处。通过跟踪磷酸化的时间进程，发现丝氨酸-587是优先磷酸化位点。还研究了蛋白C对钙调蛋白的磷酸化对肌动蛋白-H-肌球蛋白ATP酶活性抑制的影响。未磷酸化的钙调蛋白抑制ATP酶活性60%，而磷酸化的钙调蛋白几乎不抑制ATP酶活性。因此，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。
