Lin Z F, Lucero H A, Racker E
J Biol Chem. 1982 Oct 25;257(20):12153-6.
Two protein kinases (chloroplast protein kinases 1 and 2 (ChlPK1 and ChlPK2)) were isolated from spinach chloroplasts. After solubilization of the chloroplasts with octylglucoside and cholate, these kinases were purified by ammonium sulfate precipitation, sucrose gradient centrifugation, and hydroxylapatite chromatography. ChlPK1 traveled as a single band in polyacrylamide gel electrophoresis corresponding to 25,000 daltons; ChlPK2 traveled as a single band corresponding to 38,000 daltons. After exposure to 8-azido-[gamma-32P]ATP, the radioactive bands appeared in the same positions revealed by Coomassie blue staining. However, a trace of ChlPK2 was detected in the ChlPK1 preparation and a faint second lower molecular weight radioactive band was seen in the ChlPK2 preparations. Both enzymes acted on casein or histone IIIS as substrate and phosphorylated a serine residue. The proteolytic peptide maps, however, were clearly distinguishable in autoradiograms, suggesting that different serine residues were phosphorylated by ChlPK1 and ChlPK2.
从菠菜叶绿体中分离出了两种蛋白激酶（叶绿体蛋白激酶1和2（ChlPK1和ChlPK2））。在用辛基葡糖苷和胆酸盐溶解叶绿体后，通过硫酸铵沉淀、蔗糖梯度离心和羟基磷灰石色谱法对这些激酶进行了纯化。ChlPK1在聚丙烯酰胺凝胶电泳中呈现为一条对应于25,000道尔顿的单带；ChlPK2呈现为一条对应于38,000道尔顿的单带。用8-叠氮基-[γ-32P]ATP处理后，放射性条带出现在考马斯亮蓝染色显示的相同位置。然而，在ChlPK1制剂中检测到微量的ChlPK2，并且在ChlPK2制剂中看到一条较弱的第二低分子量放射性条带。两种酶都以酪蛋白或组蛋白IIIS作为底物起作用，并使一个丝氨酸残基磷酸化。然而，在放射自显影片中，蛋白水解肽图谱明显不同，这表明ChlPK1和ChlPK2磷酸化的是不同的丝氨酸残基。
