【摘要】 通过高速离心、( NH4) 2 SO4沉淀、琼脂糖凝胶过滤和 Ca M- sepharose4B亲和层析 ,从 HPGMR( 5 8S)叶片中初步纯化出 Ca2 + / Ca M依赖性蛋白激酶 .纯化的 Ca2 + / Ca M依赖性蛋白激酶经 SDS-聚丙烯酰胺凝胶线性梯度电泳 ,亚基分子量约为 5 5× 10 3.在 Ca2 + / Ca M依赖性蛋白激酶的标准反应体系中 ,存在过量的 ATP,提取磷酸化反应剩余的 ATP,用叶绿体 Mg2 + - ATP酶分解 ATP,最后用复合孔雀绿测定无机磷 ,从而来计算 Ca2 + / Ca M依赖性蛋白激酶的活性 .实验结果表明 ,该酶活性是依赖 Ca2 + 和 Ca M的 ,并且 EGTA和 TFP对酶活性有明显的抑制作用更多还原

【Abstract】 Using ammonium sulfate fraction ,sephorose CL 4B gel filtration and CaM sephorose 4B column Chromatography,a Ca 2+ /CaM PK was purified from leaves of HPGMR 58S Oryza sativa L.subsp japonica. The purified enzyme was electrophoryed by gradient SDS PAGE .The subunit molecular weight is about 55×10 3. The enzyme activity assaying system is excessive ATP in standard reaction system.The surplus ATP of phosphorylation reaction was extracted then was decomposed by chloroplast Mg 2+ ATPase. After that the inorganic phosphorus was assayed by combined malachite green reagent.The research showed that the enzyme activities are stimulated by Ca 2+ and CaM but are depressed by EGTA and TFP.更多还原