Moore J J, Moore R, Cardaman R C
Proc Soc Exp Biol Med. 1986 Jul;182(3):364-71. doi: 10.3181/00379727-182-42353.
Calcium-activated, phospholipid-dependent protein phosphorylation has not been studied in placenta. Human placental cytosol was subjected to an endogenous protein phosphorylation assay using [gamma-32P]ATP in the presence of calcium and phosphatidylserine. Protein phosphorylation was assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography. When compared to basal levels, calcium (10(-6) M) in combination with phosphatidylserine (50 micrograms/ml) significantly enhanced (P less than 0.01) 32P incorporation into phosphoproteins having mol wt 47,000, 43,000, and 37,000. Half-maximal 32P incorporation was observed with 3.5 X 10(-7) M Ca2+ in the presence of phosphatidylserine (50 micrograms/ml). The effect of phosphatidylserine was biphasic. In the presence of Ca 10(-6) M, 32P incorporation increased to a maximum at 70 micrograms/ml of phosphatidylserine. The increase was suppressed at 150 micrograms/ml. Tetracaine caused a dose-dependent inhibition of calcium-activated, phospholipid-dependent enhancement of the three phosphorproteins. Calcium in the absence of phospholipid enhanced the phosphorylation of a protein of 98,000 mol wt. Phosphatidylserine suppressed this enhancement. Calmodulin (10(-6) M) had no detectable effect upon phosphorylation beyond that of calcium alone, but the calmodulin inhibitor R-24571 specificlly inhibited the calcium-stimulated 98,000 mol wt phosphoprotein. Calcium-activated, phospholipid-dependent phosphoproteins are present in human placental cytosol; whether calcium-activated, calmodulin-dependent phosphoproteins also are present remains a question.
尚未在胎盘中研究钙激活的、磷脂依赖性蛋白磷酸化。在存在钙和磷脂酰丝氨酸的情况下，使用[γ-32P]ATP对人胎盘胞质溶胶进行内源性蛋白磷酸化测定。通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳和放射自显影评估蛋白磷酸化。与基础水平相比，钙（10^(-6) M）与磷脂酰丝氨酸（50微克/毫升）联合显著增强（P<0.01）32P掺入分子量为47,000、43,000和37,000的磷蛋白中。在磷脂酰丝氨酸（50微克/毫升）存在下，3.5×10^(-7) M Ca2+时观察到最大32P掺入量的一半。磷脂酰丝氨酸的作用是双相的。在10^(-6) M钙存在下，32P掺入量在70微克/毫升磷脂酰丝氨酸时增加到最大值。在150微克/毫升时增加受到抑制。丁卡因对三种磷蛋白的钙激活、磷脂依赖性增强有剂量依赖性抑制作用。在没有磷脂的情况下，钙增强了分子量为98,000的蛋白的磷酸化。磷脂酰丝氨酸抑制了这种增强。钙调蛋白（10^(-6) M）对磷酸化的影响除了单独钙的作用外没有可检测到的影响，但钙调蛋白抑制剂R-24571特异性抑制钙刺激的分子量为98,000的磷蛋白。人胎盘胞质溶胶中存在钙激活的、磷脂依赖性磷蛋白；钙激活的、钙调蛋白依赖性磷蛋白是否也存在仍是一个问题。
