ISOLATION AND CHARACTERIZATION OF AN ALKALINE-PHOSPHATASE FROM PEA THYLAKOIDS
ISOLATION AND CHARACTERIZATION OF AN ALKALINE-PHOSPHATASE FROM PEA THYLAKOIDS
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DOI：
10.1104/pp.99.3.1029
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发表时间：
1992-07-01
期刊：
影响因子：
7.4
通讯作者：
HIND, G
中科院分区：
文献类型：
--
作者：
KIELECZAWA, J;COUGHLAN, SJ;HIND, G
豌豆（Pisum sativum， L. cv Progress 9）光系统II叶绿素-蛋白复合物内源去磷酸化驱动状态2向状态1转变该酶是一种类囊体结合的氟敏感磷酸酶，pH为8.0时最适（Bennett J[1980]欧洲生物化学杂志，104:85-89）。从洗涤良好的类囊体中分离出具有这些特性的酶。其分子质量估计为51.5 kD，该单体具有催化活性，但活性不稳定。活性位点可用正磷酸盐在pH 5.0下标记。用测定底物4-甲基伞形草酰磷酸（350微摩尔每分钟每毫克纯化酶）获得高水平的碱性磷酸酶活性。分离得到的酶作为磷酸化蛋白磷酸酶作用于磷酸化组蛋白III-S和磷酸化的豌豆光系统ii富集颗粒，每毫克酶的典型活性为每分钟200至600皮摩尔。这些活性的最适pH值均为8.0，且对氟化物敏感。酶的最大活性需要镁离子，但不依赖于镁离子。支持该磷酸酶在类囊体蛋白去磷酸化过程中可能功能的证据来自于针对部分纯化酶的多克隆抗体制备对该过程的抑制。
Endogenous dephosphorylation of the light-harvesting chlorophyll-protein complex of photosystem II in pea (Pisum sativum, L. cv Progress 9) thylakoids drives the state 2 to state 1 transition; the responsible enzyme is a thylakoid-bound, fluoride-sensitive phosphatase with a pH optimum of 8.0 (Bennett J [1980] Eur J Biochem 104: 85-89). An enzyme with these characteristics was isolated from well-washed thylakoids. Its molecular mass was estimated at 51.5 kD, and this monomer was catalytically active, although the activity was labile. The active site could be labeled with orthophosphate at pH 5.0. High levels of alkaline phosphatase activity were obtained with the assay substrate, 4-methylumbelliferyl phosphate (350 micromoles per minute per milligram purified enzyme). The isolated enzyme functioned as a phosphoprotein phosphatase toward phosphorylated histone III-S and phosphorylated, photosystem II-enriched particles from pea, with typical activities in the range of 200 to 600 picomoles per minute per milligram enzyme. These activities all had a pH optimum of 8.0 and were fluoride sensitive. The enzyme required magnesium ion for maximal activity but was not dependent on this ion. Evidence supporting a putative function for this phosphatase in dephosphorylation of thylakoid proteins came from the inhibition of this process by a polyclonal antibody preparation raised against the partially purified enzyme.